17

Vibrio cholerae and Cholera

is probably the best known and most feared
of the diarrheal diseases discussed in this book.
Although it is by no means the most important cause of
diarrhea in terms of total morbidity or mortality, it has
caused, and in some parts of the world continues to
cause, dramatic outbreaks of acute disease accompanied by considerable loss of life. In other areas cholera
is a part of the overall spectrum of endemic diarrhea,
and in these situations it often occurs with a regular
seasonal periodicity. Cholera has a long history of
scientific investigation, with some features of its
epidemiology being clarified in London (England) by
John Snow in the 1850s; the first full accounts of its
clinical, bacteriological, and epidemiological aspects
were published in the 1880s as a result of work done in
Egypt (Koch 1884).

blood potassium levels (hypokalemia). If untreated,

some patients become rapidly dehydrated, pass into
shock, and die. Other patients experience much milder
diarrheal illness. Sixty percent or more of untreated
classical chloera cases die, whereas El Tor is generally
regarded as a milder infection with a lower fatality rate
and a higher proportion of asymptomatic infections.
Recent evidence from Bangladesh suggests, however,
that El Tor virulence may be increasing (Khan and
Shahidullah 1980). It is not possible to distinguish
classical from El Tor cholera clinically by reference to
any particular case.
The effects of cholera are due to the action of an
exotoxin, produced by the vibrios, which affects the
epithelial cells of the gastrointestinal mucosa and leads
to massive secretion of water into the lumen of the gut.
Diagnosis is by isolation of the bacteria either from
stool samples early in the clinical phase of watery
diarrhea or by rectal swab from convalescents. It is
usual to attempt direct plating on selective media as
well as enrichment in alkaline peptone water before
plating. To confirm suspected isolates, agglutination
tests with anticholera 0-group 1 serum are carried out
together with microscopic investigation for vibrio
morphology and biochemical charactrerization for
isolates failing to agglutinate. The El Tor biotype
differs from the classical vibrio in very few of its
laboratory properties.
Fatality rates can be reduced to under 1 percent in
well-managed treatment centers. The treatment of
cholera primarily consists of preventing the patient
from dying from loss of salts and water. The infection is
then self-limited, but its duration is shortened by
appropriate antibiotic therapy. Rehydration may be
by mouth in patients that are not vomiting and is by
giving clean water containing appropriate quantities of
salt, potassium chloride, alkali such as sodium
bicarbonate, and glucose to promote the absorption of
the electrolytes. Patients, particularly children, in a state
of shock or vomiting require appropriate intravenous

CHOLERA

Description of Pathogens and Disease

Despite the long history of study referred to above,
cholera is attracting renewed scientific interest, and
some traditional understandings are being considerably modified. New information is being gained
not only on the mechanisms of pathogenesis and
immunity but also on certain aspects of epidemiology
and transmission. The information summarized in this
chapter must therefore be considered as somewhat
provisional.
Identification
Cholera is caused by bacterial infection of the small
intestine. The causative organism, Vibrio cholerae,
exists in two biotypes-classical and El Tor. Both can
cause an acute intestinal disease characterized by
profuse rectal loss of water and electrolytes. The
disease begins with sudden painless evacuation from
the bowel; as it progresses, (acidotic) vomiting may
start, together with muscle cramps due to lowered

297

298

ENVIRONMENTAL BIOLOGY & EPIDEMIOLOGY: BACTERIA

fluids rapidly. Normal hydration and acid-base

balance should be achieved for adults within 2 hours of
admission to a treatment center but is achieved more
slowly for children weighing less than 20 kilograms.
Occurrence
The classical cholera vibrio is the historic cause of
cholera. From its homeland in Bengal and the Ganges
Valley, six classical cholera pandemics have spread.
The El Tor biotype, first identified in Sinai in 1905, has
only comparatively recently been accepted as V.
cholerae. A focus of El Tor cholera was known to exist
in the Indonesian island of Sulawesi in the 1930s. In
1961 this focus exploded and began to spread, thereby
initiating the seventh known pandemic of cholera. It
spread eastward to the Philippines, northward to
Taiwan and Korea, and westward into India, where it
replaced the classical biotype, and then on to Pakistan,

the Middle East, and Europe. It also spread into East

and West Africa and the Pacific islands (figure 17-1).
Where it is endemic, cholera develops a regular
periodicity, and epidemic waves occur at one or two
seasons of the year. These seasonal patterns are not the
same in various places, and there is no good
explanation of how the cholera infection cycle
correlates with climatic conditions.
Endemic cholera prior to 1960/1961 was confined to
India, especially the Ganges system, Bangladesh, and
Sulawesi. Since then it has invaded many parts of the
world and is, at the time of writing, considered to be
endemic in several areas of Africa and Asia. Many
national health authorities are very reluctant to admit
or report endemic cholera because of the possible effect
on tourism and international travel (for instance, the
pilgrimage to Mecca). For this reason endemic El Tor
cholera exists in a number of countries that officially
deny it. The present pandemic has not yet spread to the

Endemic focus of classical cholera [J

Source area of cholera El Tor
C:3

_4

First isolation of El Tor vibrios (1906)

>)

Isoloted imparted cholera cases

Figure

7-1. The global spread of chole,

5Not

/~~~~~~~~~~~~~~~~~~~~~~~~~Cp
Verd Island

\~~~~~~~~~~~~~~~~~~~~~~~~~Go
9749/7\

Figure 17-1. The global spread of cholera, 1961-75

shown
(1974)

*
I

VIBRIO CHOLERAE AND CHOLERA

Americas, although the risk of its introduction is very

great.

Injectious agents
The family Vibrionaceae includes several human
enteric pathogens of the genus Vibrio, and the
taxonomic status of some of them remains uncertain
and controversial. They are all Gram-negative, motile
rods (0.5 by 1.5-3 micrometers) usually having a
curved or comma shape. They are nonsporulating,
noncapsulated, facultative anaerobes and possess a
single polar flagellum (figure 17-2). The terminology
for the various pathogenic and closely related vibrios
used here is the one most commonly used at the present
time, although it is not ideal and may be revised (WHO
Scientific Working Group 1980).
Of greatest public health importance, and the main
topic of this chapter, are organisms that have
traditionally been called Vibrio cholerae or cholera
vibrio, but which are now strictly known as V.cholerae
0-group 1 or 01. They will be called V.cholera in this
chapter. V. cholerae is the cause of epidemic cholera
and exists in two biotypes (classical and El Tor) and
three serotypes (Inaba, Ogawa, and the much less
common Hikojima). V. cholerae produces an enterotoxin that has been extensively studied and is
similar to Escherichia coli heat-labile enterotoxin (see
chapter 13). Adherence to the intestinal mucosa is also
an important virulence factor but is poorly
understood.
A second group of V. cholerae, which agglutinate 0 1
antiserum but which do not produce enterotoxin, have

299

been recently recognized. These are known as atypical

V. cholerae 0 1 (in this chaptcr atypical V. cholerae),and
some of them have biochemical properties that differ
from those of V. cholerae. Atypical V. cholerae have
been isolated from water both in areas where endemic
clinical cholera is known to occur and in areas-such
as Brazil, England, and the USA-where it does not
occur. Atypical V. cholerae are thought not to be
enteric pathogens.
The third group of V. cholerae strains are those
which do not agglutinate O1 antisera but which are
biochemically and genetically similar to V. cholerae 01.
These are now called non-O1 V. cholerae, but until very
recently were called non-agglutinating vibrios (NAGs)
or non-cholera vibrios (NCVS). They are currently
classified into seventy-two 0-group serotypes, but this
typing scheme is tentative and provisional. Non-O1 V.
cholerae have been associated with many individual
cases of cholera-like diarrhea and with some small
outbreaks. Some non-O1 V. choleraeproduce acholeralike enterotoxin.
Finally, there are other potentially pathogenic
vibrios that are clearly not V. cholerae. V. parahaemolyticus is a halophilic marine organism responsible for numerous outbreaks and attacks of food
poisoning associated with seafood. It has a marine
rather than an enteric reservoir and so is not
considered in this chapter, although it is briefly
discussed in chapter 7. The Group F (or Group EF6)
vibrios (often mistakenly identified as Aeromonas) have
been isolated from the stools of patients with diarrhea
in many countries, but it is uncertain whether they are
toxin-producing or pathogenic. Other vibrio species

Figure 17-2. V ibrio cholerae under scanning electronmicroscopy. The single polar flagellum of the organism is
prominent. Scale bar = I micrometer. (Photo: J. Gallut, Institut Pasteur, Paris, France. Reproduced by courtesy
of Bulletin of the World Health Organization)

300

ENVIRONMENTAL BIOLOGY & EPIDEMIOLOGY: BACTERIA

occasionally isolated from man-V. alginolvticus, V.

metschnikovii, V. vulnificus, and L+ Vibrio are not
believed to cause diarrhea.
Reservoir
The primary source of infection that has been clearly
documented is the human case or carrier. There is
speculation over the role of environmental isolates of
atypical V.cholerae and non-O1 V cholerae in cholera
epidemiology and the possibility of an environmental
reservoir (see below, the section "Occurrence and
Survival in the Environment"). There is also speculation about the role of animal reservoirs, especially for
non-O1 V cholerae or for V. cholerae were isolated
interepidemic periods. Sanyal and others (1974)
examined 1,287 fecal samples from 195 domestic
animals following an outbreak of cholera in Varanasi
(India) during 1972. The proportions of animals from
which V. cholerae or non-O1 V.cholerae were isolated
were: dogs, 27 percent; chickens, 18 percent; cows and
goats, each 11 percent. There were no isolations from
buffalo, donkeys, or horses. Out of a total of fifty-four
strains of V-cholerae isolated, eight were V cholerae 01
(El Tor, Ogawa). Neither this nor other studies have
clearly shown that animal infections with V.cholerae or
non-O1 V. cholerae play any role in the epidemiology of
human infection and disease.

Cholera is transmitted by the fecal-oral route from

person to person, and transmission is encouraged by
inadequate water supply and excreta disposal facilities
b gneralv,
povrty ad ovrcrowing.
and, ore
and moreseneraly 'by mptovet andiovercro mi
Convalescent and asymptomatic individuals may
excrete 10' 105 V. cholerae per gram of feces, whereas
an active case excretes
V10e 10
per milliliter of ricewater stool (Dizon and others 1967; Greig 1914;
Smith, Freter and Sweeney 1961).
Infective doses are high in healthy adult males.
Hornick and others (1971) required 108 classical V.
cholerae in water to produce diarrhea in 50 percent of
adult volunteers (the median infective dose, or ID5 0 ),
and 10" organisms to produce cholera-like diarrhea.
With the prior administration of 2 grams of sodium
bicarbonate, the ID 5 0 was lowered to 104 for diarrhea
and 108 for cholera-like diarrhea. No diarrhea or
infection was produced by < 108 organisms without
NaHCO3 or by < 103 organisms with NaHCO3 (see
also Cash and others 1974).
Gastric acidity is an important barrier to cholera
infection, and those with lowered acidity (hypo-

chlorhydria) may be infected by lower doses than

others. More recent volunteer studies with El Tor
strains have shown that infective doses are lower when
the organisms are administered in food than in small
volumes of water (WHO Scientific Working Group
1980). This could be due to more rapid gastric
emptying, neutralization of gastric acid by food, or
protection of vibrios that are adsorbed to, or
embedded within. food particles. Nothing is known
about the dose needed to cause acute diarrhea in 1
percent of malnourished children, but it may be 102 or
even less.
If it is assumed that the environmental reservoirs of
V. cholerae described below are epidemiologically
unimportant, then cholera transmission must take
place by direct person-to-person contact or by the fecal
contamination of water or food. Waterborne and
foodborne transmission have both been clearly
demonstrated on specific occasions. Cholera has
classically been regarded as a waterborne disease, and
there are some experts who believe that this is its
dominant and normal mode of transmission. Others
maintain that this may be true in Bangladesh but not
elsewhere, while a third opinion holds that cholera
transmission among poor people in developing
countries is primarily nonwaterborne. This subject has
attracted recent debate (for instance Feachem 1976;
Levine and Nalin 1976) and is of considerable
importance in designing control strategies. The topic
has been comprehensively reviewed bv Feachem
(1981. 1982).
Incubation period
The incubation period is generally short and clinical
symptoms occur within 0.5 to 5 days (usually 1-3 days)
o netn h atra
nuainprosmyb
of ingesting the bacteria. Incubation periods may be
inversely related to the dose of organisms ingested.
Period of communicability
Convalescents generally excrete V. cholerae intermittently and only for short periods. Thus, 50 percent
of cholera cases will be found to excrete the pathogen
for up to 5 days, 30 percent continue to excrete for up to
15 days, and 10 percent for up to 25 days. By 1 month
usually less than 5 percent of cases are still excreting V.
cholerae, and it is very uncommon to find carriage
persisting beyond 2 months. The truly chronic
carrier such as Cholera Dolores from the Philippines
(Azurin and others 1967)-is a very rare phenomenon.
Asymptomatic infection is common, and the El Tor
biotype produces a higher infection to case ratio than
classical cholera.

VIBRIO CHOLERAiE AND CHOLERA

301

Resistance

plausible but usually unproven explanations of

In edemcapeas
aras,it tht rpeatd rinwaterbornc transmission.
fcion endemichareas, itapears thatgrepeate reind-upo One of the most characteristic features of endemic
immunity with increasing age (Gangarosa and Mosley
1974). This may be one reason why the attack rates in
children in endemic areas are considerably higher than
in adults, whereas in epidemic situations where cholera
has been recently introduced the reverse is often true.
However, among those infected overt disease is more
common in adults than in children.
A previous attack of cholera diarrhea confers solid
immunity against reinfection with the same serotype of
V. coleae
bou Ior ear.An nvetigtio in
VBahorain
fhore tabot infyear.wAn investigationlin
bahrai-fe showd signifiantsy
tat
whogwere prinkofcipally

instance, int Dacca (Banonglds)choeaued

sesnlpttmForpa
dramatc,ic allyaduringlNovemb
oer-anuary,
whereask0
kilometersaway duing Calcuttr-aIndiarythereak was
klmtrawy
i
acta(ni)tepa
a
April-June. Recently these peaks have shifted and
no ocudrigSpeb-Nvmrin
ot
areas. The reasons for these and other seasonal
patterns of cholera remain entirely unexplained.
Non-0l V. cholerae has been isolated from stools of
persons with diarrhea in many countries in Asia,
Africa, Europe, and, significantly, North and South
America. Large epidemics have not been reported. In

than infants who were breast-fed, although it was not

clea arse
whtherom
thi ontminted ilkand
cotlear whthrtirs from
nreiitaminmatedna
conciv
milkan
bottesno from pothectiv inreint7n9aeraml

thUS motifconocudrnghewmr
summer months, while in Bangladesh there appears to
be a peak in spring and summer before the annual
cholera peak. Small foodborne outbreaks are common

Cholera is a disease of the lower socioeconomic

groups. Fishermen and boatmen, living along polluted
water courses, are specially at risk. So also are people
with hypochlorhydria, either due to malnutrition or
other natural causes, or following gastric surgery (Sack
and others 1972). Although the El Tor biotype may be
less virulent than the classical, causing more mild cases
of cholera, the host is probably equally susceptible to
colonization by either.

intransmission e
onre,btltlskono
cutrasies.in adeieilg
ndvlpn
Thenepidemilgsfcolr.ean i
aywy
unc ertinademontovyocoersal Theman inmprance ofy
ucranadcnrvril
h
motneo
waterborne transmission, the maintenance of cholera
during interepidemic months of the year, the
explanation of seasonality, the failure of tubewells in
Bangladesh to reduce incidence, and the role of a
possible aquatic reservoir for V cholerae are all
topics of current debate. Space does not permit a full
~~~~~~~~review
of these issues here. For a conventional account
~~~~~~~of
cholera epidemiology, the reader should consult
Gangarosa and Mosley (1974); Feachem (1981, 1982)
provides a review of the more recent literature and
debates.

Epidemiology
Epidemiology
Studies on V. cholerae El Tor infection, in both
epidemic and endemic situations, have repeatedly
emphasized that the severe eases that reach the
attention of treatment centers and physicians are the
tip of an iceberg of widespread asymptomatic and mild
clinical infection in the community. Estimates of a case
to infection ratio of 1: 30, or less, are commonly quoted.
The asymptomatic infections are generally short lived
but can be of crucial epidemiological importance in
transmitting and geographically spreading cholera.
Attempts to reconstruct the modes of transmission and
spread of cholera that concentrate on known clinical
cases are unlikely to be successful. To understand
cholera epidemiology, it is necessary to take full
account of the transient carrier, and to document the
occurrence of transient carriage it may be necessary to
undertake multiple fecal examinations and use
serological techniques to determine whether an
asymptomatic individual has been infected. These
difficulties are one reason why so many investigations
of cholera outbreaks are inconclusive or fall back on

Control Measures
The most cost-effective control measures to deal
with either endemic or epidemic cholera remain
uncertain. Understanding of control will increase as
more information is gathered on the epidemiological
issues discussed above. Cholera control among people
who are poor has so far proved to be extremely difficult.
The course of a cholera epidemic is often dramatic and
short-lived, and by the time control measures are
applied the epidemic may be waning naturally. This
can give a false impression of the efficacy of the control
measures and lead to unjustified claims--as was the
ease when John Snow removed the handle from the
Broad Street pump in London (England) in 1855.

302

ENVIRONMENTAL BIOLOGY & EPIDEMIOLOGY: BACTERIA

Individual

have been made for the efficacy of various environmental control methods, but few of these have
been justified, and most programs have been
unsuccessful. Indeed, the experience with environmental control among the rural and urban poor has been so
bad that some experts feel that the priority allocation
of resources should be toward the establishment of

Prophylactic antibiotics have been used to control

some cholera outbreaks and to limit their spread.
There is no evidence that this practice is effective, and
there is mounting concern over the rising prevalence of
antibiotic-resistant strains of V. cholerae in some
countrles. Large amounts of tetracycllne ( 1,788
kilountries. in e firta6mounths)of tetracyclined the
i
networks of treatment centers for providing simple but
kilograms in the first 6 months) were used therapeutihighly effective rehydration therapy to reduce morte e
higy (efectiv
e
rrp
cally and prophylactically following the outbreak of
all
atY
(Greogh
17).
cholera in Tanzania in October 1977. Initially,
chola in T a iThe impact of water supply and sanitation schemes
.o.n.i.i
strains of V. cholerae tested were fully sensitive to
bu afte 6 moth 76pretofioae
on endemic or epidemic cholera in poor communities iS
tetracycline, but after 6 months 76 percent of isolates
uncertain. Six studies in Bangladesh showed no impact
were resistant (Mhalu, Mmari and ljumba 1979).
(Briscoe 1978; Feachem 1982), whereas a study in the
Subsequent work showed that this antibiotic resistance
P
was mediated by transferable plasmids that confer
aPhilippines showed a very considerable impact (Azurin
multiple antibiotic resistance (Towner and others
and Alvero 1974) Thesnterpretatlon ofthese findgs
Tonr1ero
99.Mlil
~~,Multiple
S controversial and has been recently reviewed in
1980;~
~Pearson
~ and
n ~ O'Grady
'rd~
~1979).
1980; Towner,
dealbFece(18)
antibiotic resistance has also been reported from 5-36
percent of V. cholerae isolates from Bangladesh
In some outbreaks-for instance, in Tanzania from
(Threlfall, Rowe and Huq 1980).
1977 to 1980-the geographical spread of cholera was
.
due to the movement of infected individuals and gave
t
8) .
(Thefl,owclpeeand
rise to the characteristic pattern of spread along major
isaffd
at psresn
dimmupoinolog.Kicalpeventio vaccines
disappointing. Killled vaccines do afford a measure of
ala
n
odrue.
nsc
icmtne
h
protection but are usually less than 70 percent effective,
7 poerenot elcte
protectionmbuntyar usually less t
and such immunity as IS produced does not last at

railway and road routes. In such circumstances the

reasonable levels for more than about 4 months. A

study In Bangladesh Sowed
ass vaccinaion3wa

known to be affected may reduce the risk of spreading

the disease. Travel restrictions are difficult to enforce,
however, and may seriously disrupt the movement of

Current
neffected( tfmm
er
Mosceyi173)
cractdering
Currogentesesarch isvirlented factor
fut
pathogenesis and virulence factors and at developing
and testing a variety of alternate vaccines based on
live mutant strains or nonviable antigens such as the B
subunit of the cholera enterotoxin.
Rigorous personal cleanliness and care in eating and
drinking habits are probably the surest ways by which
an individual
can reduce the risk of cholera in an

foodstuffs. If travel restrictions are combined with

issuing prophylactic tetracycline to those who must
tae,a
a
oei
azna h
rbeso
travel, as was done in Tanzania, the problems of
increased antibiotic resistance described above may
occur.
Cvjetanovid (1979) and Cvjetanovi', Grab and
Uemura (1978) used a mathematical model to compute
the relative economic merits of sanitation, chemopro-

endemic or epidemic situation.

phylaxis, and immunization as methods of cholera

control. Unfortunately, the cost of sanitation was set
far too low (US$0.15 per capita at 1971 prices), and the
effectiveness of sanitation was overestimated. Not
surprisingly, this analysis showed sanitation to be
highly cost-beneficial (with benefits taken only as the
medical treatment costs saved), whereas immunization
was shown to have costs far exceeding benefits because
the currently available vaccine would have to have
been given annually to have had any major impact on
disease. Nonetheless, the analysis highlighted the
benefits of sanitation as a measure having potential
effects on a range of enteric and other diseases, as
compared with vaccination, which, even if a more
protective vaccine were available, is difficult to
administer to most children, probably requires
repeated readministration, and only protects against a
single pathogen.

Environmental
There is no doubt that some combination of
improved water supplies, excreta disposal facilities,
better housing, and all the various improvements in
daily life that come with increased wealth and
education have been responsible for the elimination of
cholera from the developed countries and from many
middle-class communities in developing countries.
Cholera was and remains a disease of poverty and the
living conditions that are associated with poverty.
Countries that experience the problem of endemic or
epidemic cholera today are faced with the question of
how to control the disease among poor communities in
the short-term while poverty persists. Many claims

limitation of movement of people in or out of areas

VIBRIO CHOLERAE AND CHOLERA

Carrier surveillance and internationalregulations

Since the chronic carrier is extremely rare,
surveillance to identify carriers is notof significance i
the control of this disease. This is in m arked contrast
with typhoid. The principal types of cholera carriage are
icubatory, convalescent, and contact.
Up to December 31, 1970 International Sanitary
Reguatios
i fore.
wre hey tipuate a 5day
Regulations were in
force. They stipulated a 5-day
quarantine
speriod.for
relesfom areasahr
cholera was established. The regulations were abandoned when it was recognized that they were not
preventing the spread of the current pandemic. Among
the reasons for this failure were the concealment or
denial of the existence of the disease in a country,
together with the unknown importation of cases across
unpatrolled borders. Current surveillance at national
udnpaterolledtbonaldleversCurrent svenianfcati
in
and international levels has been ineffective
preventing the spread of cholera into receptive
countries-those with poor sanitation, hygiene, and
health services. Nonetheless, surveillance to identify
clinical cases (and, hence, the geographical advance of
the disease) provides valuable epidemiologcal
minformation and allowsthe organizationoftreatmentinthe
absence of effective control measures.
Occurrence and
Environment

Survival in the

The study of V cho*erae, atypical V cholerae, and

non-01 V cholerae in the environment is attracting
increasing attention at the present time. The conventional view that V. cholerae is an organism only
found in the environment in close association with
human cases or infections, and only surviving for a few
days at most, is now being revised.

The relationship between V. cholerae and water has

been the focus of many investigations and is crucial to
an understanding of the epidemiology of cholera. The
traditional view of this subject-as stated by Felsenfeld
(1974):
some authors claimed that cholera vibrios may
survive in water, particularly, seawater, for as long as
2 months. This is, however, scarcely possible under
natural
' conditions
reinfection of te w r d
natural conditions if reinfection of the water does
-is

now known to be incorrect.

Data on the occurrence of V. cholerae in water are of
two types. First, there are the numerous reports of V.

303

cholerae isolations from rivers, tanks, ponds, wells, and

household water jars in or near communities where
cholera cases or infections are known to be occurring.
Some of these reports are reviewed in a separate
publication s(Feachems1981) eSecond theresarerte
publication (Feachem 1981). Second, there are the
more recent findings of V. cholerae, especially but not
exclusively atypical 01 and non-Ot strains, in water
an watwae atstsdsatfo
n nw ua
and wastewater at sites distant from any known human
V choleraeinfection. These findings are reviewed below
in the section on possible aquatic reservoirs.
The reason that the view expressed by Felsenfeld was
so strongly held for nearly 100 years is, first, that
researchers had failed to find V cholerae in the aquatic
environment except in close association with human
neto det
obnto
fntloig
okn
infection (due to a combiation of not lookmg, loohng
in the wrong manner and looking in the wrong place),
and, second, that survival experiments conducted in the
laboratory had shown V cholerae to be an organism
with only limited survival ability in certain aquatic
environments.
Some of the considerable accumulation of data on V.
cholerae survival in water is summarized in tables 17-1
to 17-5. In clean water (for instance, dechlorinated tap
water), survival times are up to 1 month at 4C and
2-14 days at 20-30C. In raw well water, survival times
are over a month at 4C and generally between 1 and 20
days at 20-30C, although reports from India and
Tanzania suggest survival of the El Tor biotype in raw
well water of up to 55 days. A single report of V.
cholerae survival in refrigerated raw surface water gives
a survival time of 48 days, while survival at 20-30'C is
generally 1-6 days, with occasional reportings of
longer survival and one exceptional report from
Tanzania of 48 days. As would be expected, survival in
seawater is prolonged, with durations of 2 months at
4XC and 6-60 days at 20-30'C. Finally, a single report
from the USSR (table 17-4) and epidemiological
evidence from Portugal (Blake and others 1977)
suggest the ability of V. cholerae to survive for
prolonged periods in certain mineral waters.
It is clear from the tables that survival can be greatly
prolonged in nutrient-rich waters and seawaters that
have been boiled or autoclaved prior to contamination
with V cholerae, thus eliminating competing microorganisms and possibly also making the chemical
comp osition of the water more favorable
for V
chosition o
the
u
nature avorable or V
choleraesurvival. Although the nature and extent of V.
cholerae inhibition by a mixed microflora in a natural
surface water are not known, one study showed a
failure of E. coli, Pseudomonas spp., and Aerobacter
spp. to suppress V. cholerae El Tor survival in artificial
sterile well water (Pandit and others 1967). Sunlight
considerably curtails V.cholerae survival.

304

ENVIRONMENTAL BIOLOGY & EPIDEMIOLOGY: BACTERIA

Table 17-1. Survival of Vibrio cholerae in surface waters

Source

Biotype and
initial concentration
per milliliter

Type of sample

Temperature

Cheng (1963)

El Tor
1.5 x 105

River water
Drain water
Pond water
(all taken in
or near Taipei)

21-31C

Gohar and
Makkawi
(1948)

Classical
from feces
from culture

Nile water

Room temp.
(Egypt)

Khan and
Agarwal
(1929)

Classical
(clinical isolate)

Jumna and Ganges

river waters
Raw
Filtered
Boiled
Boiled & filtered

Non-Of
(water isolate)

3 days
2 days
6 hours

5 days
10 days

Room temp.
(Allahabad)
8 days
18 days
29 days
14 days

Raw
Filtered
Boiled
Boiled & filtered

20
20
18
20

Konchady
and others
(1969)

Classical
104

Calcutta
River Hooghly
Canal water
Pond water

250 C

Lahiri, Das
and Malik (1939)

Classical
(Inaba)

Spring water
Raw
Autoclaved

Room temp.
(Calcutta)

106

Survivala

days
days
days
days

6 days
6 days
6 days

1 hour
18 hours

River Hooghly
(Calcutta)
Raw
Autoclaved
Filtered
Autoclaved &
fiitered

18 hours
3 days
2 days

Tank waters
(Calcutta)
Raw
Autoclaved
Filtered
Autoclaved &
filtered

2 3 days
3-12 days
7 days

2 days

15-18 days

Lema, Ogwa
and Mhalu
(1979)

El Tor
105

Swamp water in Dar

es Salaam

4C
30C
32C in
sunlight

Mukerjee, Rudra
and Roy (1961)

Classical
2 x 106

River Hooghly
(Calcutta)

Room temp.
(Calcutta)

48 days
48 days
3 days

Raw

1-6 days

Autoclaved
Filtered

4-22 days
3-12 days

305

VIBRIO CHOLERAE AND CHOLERA

Table 17-1 (continued)

Source

Biotype and
initial concentration
per milliliter

El Tor
(clinical isolate)
2 x 106

El Tor
(water isolate)
2 x 106

Non-OI
(clinical isolate)
2 x 106

Non-O1
(water isolate)
2 x 106

Type of sample

Temperature

SurvivaP

Tank water
(Calcutta)
Raw
Autoclaved
Filtered

1-6 days
4-23 days
3-7 days

River Hooghly
(Calcutta)
Raw
Autoclaved

2 days
11 days

Tank water (Calcutta)

Raw
Autoclaved

2 days
13 days

River Hooghly (Calcutta)

Raw
Autoclaved

2 days
11 days

Tank water (Calcutta)

Raw
Autoclaved

2 days
16 days

River Hooghly (Calcutta)

Raw
Autoclaved

2 days
9 days

Tank water (Calcutta)

Raw
Autoclaved

2 days
12 days

River Hooghly (Calcutta)

Raw
Autoclaved

2 days
11 days

Tank water (Calcutta)

Raw
Autoclaved

2 days
13 days

Neogy (t965)

Classical
El Tor

Pond water

Room temp.
(India)

1-2 days
8 days

Read and
others (1939)

Classical

Autoclaved tank
waters (Calcutta)

Room temp.
(Calcutta)

>30 days

Note: Older literature is reviewed by Pollitzer (1959).

a. Times given, for instance, as 22 days are durations at which viable organisms could no longer be detected. Times given as > 30 days
indicate that organisms were still viable at that time but that sampling was discontinued.

Some experiments have included direct comparisons

of the survival of classical and El Tor biotypes, and
occasionally also non-O1 strains (tables 17-1, 17-2 and
17-4). Two studies showed markedly longer survival of
El Tor than classical V. cholerae (Felsenfeld 1965;
Neogy 1965); one study showed similar survival
between the two biotypes (Sayamov and Zaidenov
1978); one study showed non-O1 V. cholerae surviving

for longer than classical V. cholerae 01 (Khan and

Agarwal 1929); and one study showed no difference in
survival between classical O1,ElTorOl and non-O1 V.
cholerae (Mukerjee, Rudra and Roy 1961). It would
appear from this literature review that the widely held
belief that El Tor V. choleraesurvives for considerably
longer periods in water than the classical biotype is not
firmly based. This is especially true in view of the major

306

ENVIRONMENTAL BIOLOGY & EPIDEMIOLOGY: BACTERIA

Table 17-2. Survival of V. cholerae in well water

Biotype and
initial concentration
per milliliter

Source

Type of sample

Temperature

Survivala

Cheng
(1963)

El Tor
1.5 x 105

Well water
(village near Taipei)

21-31C

I day

Felsenfeld
(1965)

Classical
El Tor

Shallow well water

?
?

8 days
19 days

Khan and
Agarwal
(1929)

Classical
(clinical
isolate)

Well water
(Allahabad)
Raw
Filtered
Boiled
Boiled & filtered

Room temp.
(Allahabad)

Non-OI
(water
isolate)

Raw
Filtered
Boiled
Boiled &filtered

Konchady
and others
(1969)

Classical

Lema, Ogwa
and Mhalu
(1979)

McFeters
and others

I
6
9
8

day
days
days
days

12 days
6 days
18 days
26 days

Well water
(Calcutta slum)

25C

6 days

El Tor
105

Well water
(Tanzania)

4C
30C
320 C in
sunlight

55 days
55 days
I day

Sterile well water

9.5-12.5C

>2 days

104

105

(tso = 1.3 days)b

(1974)
Pandit and
others
(1967)

El Tor
(Ogawa)

Well water
(Punjab)

21C
37C

18 days
4 days

Well water
(Uttar Pradesh)

2 VC
250 C

37C

51 days
Fourfold growth
after 1 day
Survival for
> 7 days
4 days

25C

10-12 days

5-100 C
30-32C
Sunlight
5-10C
30-32C
Sunlight

18 days
13 days
4 days
42 days
17 days
8 days

103

Experiments with well water

simulating actual removal
and replacement of water
in well following single
contamination with 103 V
cholerae per milliliter
Pesigan,
Plantilla
and Rolda
(1967)

El Tor
106

Deep well water

(Manila)
Raw

Autoclaved

307

VIBRIO CHOLERAE AND CHOLERA

Table 17-2 (continued)

Source

Biotype and
initial concentration
per milliliter

Type of sample
Raw well water stored
in clay jar

Shrewsbury
and
Barson
(1957)

Classical

30-32C
ambient, but
jar storage
may have
cooled water

Sterile, synthetic
well water of same
composition (pH = 5.6)
as Hagar's Well
(Mecca, Saudi Arabia)
during the cholera
epidemic of 1883
Same water with:
pH 7
pH 8
pH 9

Temperature

Survivala
32 days

5C
21C
25CC

1 day
I day
I day

5C
21'C
5C
21C
5C
21C

3 days
3 days
3 days
77 days
3 days
3 days

Note: Older literature is reviewed by Pollitzer (1959).

a. Times given, for instance, as, 18 days are durations at which viable organisms could no longer be detected. Times given as > 7 days indicate
that organisms were still viable at that time but that sampling was discontinued.
b. t55 Time for 90 percent reduction.

probable strain-by-strain differences within each

biotype and the differences between laboratory
cultures, fresh clinical isolates, and water isolates. On
the basis of the literature reviewed here it remains
unproven than El Tor is a more persistent organism in
water than the classical biotype, and the true
interbiotypic and intrabiotypic variabilities in survival
remain to be documented. It follows that explanations
of the differences in epidemiology between El Tor and
classical cholera-for instance, the greater "endemic
tendency" of the former-cannot, at the present time,
make use of putative differences in environmental
persistence between the two biotypes.
Laboratory experiments on V. cholerae survival in
water may accurately reflect conditions in manmade
containers of clean water (such as reservoirs, cisterns,
jars, and glasses), but they cannot replicate conditions
in natural water bodies such as rivers, ponds, or even
open wells. In these latter waters there may be
abundant flora and fauna, and many varied surfaces,
not reproduced or simulated in the laboratory
experiments. There is increasing evidence (reviewed
below) that V. choleraein natural waters are frequently
in close association with bottom sediments, chitinous

fauna, and plant surfaces; therefore, laboratory data

must be interpreted with extreme caution.

Fxcept for the atypical V cholerae and non-O1 V.

cholerae which may maintain an environmental
reservoir, the primary source of V. cholerae in the
environment is the feces of man. Persons infected by V.
cholerae, though not sick, may excrete 102-105 per
gram of feces, while those with active and severe disease
may excrete 106-109 per milliliter of rice-water stool
(Dizon and others 1967; Greig 1914; Smith, Freter and
Sweeney 1961). Unlike most other enteric bacterial
infections, the prevalence of excretion of V. cholerae by
the general healthy population is very low-typically
well under 1 percent, even in endemic areas.
In areas of endemic cholera, or during a cholera
outbreak, it is to be expected that V. cholerae will occur
in the night soil produced by the affected communities.
Forbes, Lockhart and Bowman (1967) and van de
Linde and Forbes (1965) reported numerous isolations
of V. cholerae from night soil in Hong Kong, both when
cholera cases were and were not occurring in the city.

308

ENVIRONMENTAL BIOLOGY &EPIDEMIOLOGY: BACTERIA

Table 17-3

Survival of V. cholerae in tap water

Source

Biotype and
initial concentration
per mnilliliter

Type of samnple

Cilt/rine reSidL1l
milligrams
per liter

Temperatu7e

Surcirala

Cheng
(1963)

El Tor
1.5 x 105

Taipei tap
water

21-31CC

0.5

2 hours

Konchady
and others
(1969)

Classical
104

Tap water
from deep
tubewell
(Calcutta)

25C

6 days

Lahiri, Das
and Malik
(1939)

Classical
(Inaba)
106

Calcutta tap
water
Raw
Autoclaved
Filtered
Filtered &
autoclaved

Room temp.
(Calcutta)

?
18 hours
24 hours
2 days
12 days

Lema, Ogwa
and Mhalu
(1979)

El Tor
105

Dar es Salaam
tap water

4C
30C
32C in
sunlight

Chlorinated
at treatment
works but
probably no
residual
chlorine
remaining
at tap

34 days
14 days
3 days

Mukerjee,
Rudra and
Roy
(1961)

Classical
2 x 10'

Calcutta tap
water
Raw
Autoclaved
Filtered

Room temp.
(Calcutta)

Pandit and
others (1967)

El Tor
(Ogawa)

Delhi tap
water

21C
37C

De-chlorinated

12 days
1 day

Manilla tap
water
Raw
Raw
Raw
Autoclaved
Autoclaved
Autoclaved

5-1 0C
30-32C
Sunlight
5- l0C
30-32C
Sunlight

0.6
0.6
0.6
0
0
0

1 hour
1 hour
1 hour
10 days
1.6 days
12 hours

2-8 days
4-18 days
2-6 days

to3

Pesigan,
Plantilla
and Rolda
1967)

El Tor
lob

Note: Older literature is reviewed by Pollitzer (1959).

a. Times given are durations at which viable organisms could no longer be detected.

During a 10-month sampling period, 46 percent (200 of

433) of bucket latrines in the slums of eastern Calcutta
(India) were positive for V. cltolerae on one or more
occasions (Sinha and others 1967). V. cholerae
isolations from latrines were obtained during months
when no cholera cases were reported. In contrast,
during 1968 in Dacca and Chittagong (Bangladesh) a

total of 72.494 night soil samples yielded only 56

isolations of V.cholerae. all of which occurred at times
when cholera cases were being reported (Bart, Khan
and Mosley 1970).
Some reported data on V. cholerae survival in feces
are summarized in table 17-6. Clearly survival is
inversely related to temperature. Cheng (1963) and

l'IBRIO

309

CHOLERAE AND CHOLERA

Table 17-4. Survival of V. cholerae in mineral water

Source
Sayamov and
Zaidenov
(1978)

Biotype and
initial concenitration
per milliliter
Classical
9 x 105
1.5 x 103
9.5 x 10W
1.6 x 10'
El Tor
1.2 x 106
t03
9 x lo,
1.6 x 10'

Type of samnple
Spring water from spa
(Matsesta, USSR)
Raw
Diluted
Boiled
Diluted
Raw
Diluted
Boiled
Diluted

Temperature

20-24C
370C
20-24C

37C

Survivala

22
15-65
> 1429
>289

days
days
days
days

22 days
18-39 days
>1429 days
>413 days

Note: Further evidence of prolonged survival of V cholerae in mineral water is provided by the investigation of the cholera outbreak in
Portugal in 1974 (Blake and others 1977).
a. Times given, for instance, as 22 days are durations at which viable organisms could no longer be detected. Times given as > 289 days
indicate that organisms were still viable at that time but that sampling was discontinued.

Shoda, Koreyeda and Otomo (1934) found that

survival was longer in liquid stools than in soft or solid
stools. In summary, at ambient temperatures in tropical
and subtropical countries, V. cholerae is unlikely to
survive beyond 5 days in feces.
In sewage
There are very few reports of V. cholerae in sewage.
This is primarily because, in most developing
countries, the section of the population that experiences the highest attack rates of cholera produces
no sewage because their houses do not have flush
toilets. Instead, they produce night soil (where V.
cholerae has been found) or they defecate beside or
into open water bodies (where V. cholerae has also
been found).
Kott and Betzer (1972) reported estimates that
Jerusalem sewage contained between 10 and 104 V.
cholerae per 100 milliliters during the 1970 cholera
epidemic in Israel. Daniel and Lloyd (1980a) reported
geometric mean concentrations of 2,600 and 160 non01 V. cholerae per 100 milliliters of very strong sewage
(suspended solids 17,000 and 7,400 milligrams per liter,
respectively) in two refugee camps near Dacca
(Bangladesh). Isaacson and others (1974) reported the

use of Moore pads to detect V.cholerae in sewage at

mines in the Transvaal (South Africa) during 1973-74,
when the spread of cholera from Malawi,
Mozambique, and Angola was feared. V. cholerae (El
Tor, Inaba) was isolated from the sewage prior to and
during cholera outbreaks at the mines and acted as an
effective early warning system for the outbreaks.
Survival of V. cholerae in sewage is summarized in
table 17-7. Three studies (Altukhov and others 1975;

Daniel and Lloyd 1980b; Zaidenov and others 1976)

suggested that some sewages provide a permanent
culture medium for some strains of classical, El Tor,
and non-O1 V. cholerae. The other studies found that
survival times were 1-24 days in sewage at 20-30C.
Survival times are shorter at warmer temperatures and
longer in sterilized sewage than in raw sewage.
Direct comparisons of different biotypes and
serotypes showed no differences in survival among
classicalO1, El Tor 01,and non-Ol strains (Mukerjee,
Rudra and Roy 1961). Altukhov and others (1975)
found an El Tor, Ogawa strain better able to multiply
in bath house sewage at 37C than a classical, Ogawa
strain, although even the classical strain had not fallen
below its initial concentration after 10 days. Daniel and
Lloyd (1980b) found a sewage-derived non-01 strain
better able to multiply in sewage than a laboratory
reference strain of El Tor 01, although even the El Tor
strain showed no reduction in concentration between 6
hours and 48 hours at 22-25C. As with water,
therefore, there is little evidence at present to suggest
that the El Tor biotype is necessarily better able to
survive in sewage than the classical biotype.

Summary of survival in water and wastewater

In some survival studies the initial concentration of

organisms present was reported, and it is therefore
possible to estimate a death rate expressed as a tgo
value-the time in hours for a 90 percent or 1 log unit
decline in concentration. In only a few studies were
death curves plotted from which accurate tgo values
might be taken. For other studies the tgo value can only
be estimated from the initial concentration and the

310

ENVIRONMENTAL BIOLOGY &EPIDEMIOLOGY: BACTERIA

Table 17-5. Survival of V. cholerae in seawater

Source

Biotype and
initial concentration
per milliliter

Type of sample

Cheng (1963)

El Tor
1.5 x 105

Coastal water
near a freshwater source

Jamieson,
Madri and
Claus
(1976)

El Tor
1.5 x 107

Sterilized seawater
with adjusted salinity (percent)
0.5

2.0

3.5

Lema, Ogwa
and Mhalu
(1979)

El Tor

Pesigan,
Plantilla
and Rolda
(1967)
Various
studies
between
1885 and
1920
reviewed
by Pollitzer

Temperature

Suirvirala

21-31C

6 days

4C
25C
37C
4C
25C
37C

5 days
3 days
2 days
4 days
3 days
1 day

4C

4 days

250 C
37C

1 day
1 day

Seawater
(Dar es Salaam)

4C
30C
320 C in
sunlight

> 58 days
>58 days
5 days

El Tor
106

Seawater (Manilla)

5-10C
30 32C
Sunlight

58-60 days
10-13 days
10- 1 days

Classical

Sterilized seawater
(Marseilles)
Seawater
(Copenhagen)
Seawater (New York)
Raw
Sterilized

81 days

Summer
Winter

7-17 days
47 days

?
?

7-47 days
>285 days

Seawater (Japan)
Raw
Raw
Raw
Sterilized
Sterilized
Sterilized

4C
Room temp.
37C
4C
Room temp.
37C

9-27 days
7-41 days
3-12 days
53-230 days
152 209 days
30-83 days

Artificial seawater
Top of tank
Bottom of tank
In sunlight

18C
18C
19-40C

23 days
30 days
2 hours

105

(t959j

Yasukawa
(1933)

Classical
3 x 104
3 x 105

a. Times given, for instance, as 6 days are durations at which viable organisms could no longer be detected. Times given as > 58 days indicate
that organisms were still viable at that time but that sampling was discontinued.

311

VIBRIO CHOLERAE AND CHOLERA

Table 17-6. Survival of V. cholerae in feces

Source

Biotype and
initial concentration
per milliliter

Type of sample

Temperature

Survivala

Abel and
Claussen
(1895); cited
by Pollitzer
(1959)

Classical

Naturally infected
cholera stools

13 16CC

10 days
for over
half the
samples with
a maximum of
29 days

Cheng
(1963)

El Tor

Naturally infected
stools
Artificially
infected stools

29-31C

1-4 days

29-31C

2 4 days

Gildemeister
and
Baerthlein
(1915); cited
by Pollitzer
(1959)

Classical

Naturally infected
stools

12-21C

10 days for
half the
samples;
with a
maximum of
51 days

Greig
(1914)

Classical
1.5 x 1082 x 109

Naturally infected
ricewater stools

22C

Min. 1-3 days

Max. 10-17 days
Av. 3-8 days
Min. 1 day
Max. 2 13 days
Av. 1-7 days

29C

Shoda,
Koreyeda and
Otomo (1934)

Classical

Naturally and
artificially infected
stools

a. Times given are durations at which viable organisms could no longer be detected. Max.

overall survival time, without knowing the shape of the

intervening death curve or whether the number of
organisms fell below detectable levels considerably
prior to the stated survival time.
Bearing in mind these limitations, tgo values have
been derived where possible. The few studies that
showed prolonged maintenance of concentrations
equal to or greater than initial values have been
excluded and are discussed separately in the next
section. Derived tgo values are presented in table 17-8.
The mean figures in table 17-8 suggest maximum
survival in well water and seawater. The mean figures
for the El Tor biotype are greater than for the classical
biotype, but this comparison is invalid since each
experiment used very different techniques and a wide
variety of strains of various origins. It remains

4C
Room temp.
(Japan)
37C
=

1-5 days
0.5-2 days
6 hours

maximum, Min. = minimum, Av. = average.

uncertain whether the interbiotypic variability of

survival is greater than the intrabiotypic variability.
These tgo values may be compared with typical tgo
values for coliforms of 20 to 115 hours (median 60
hours) in surface waters and with 0.6 to 8 hours (mean 2
hours) in seawater (chapter 13). For shigellac, in
surface waters at temperatures of over 20C, tgo values
generally fall well below 60 hours (chapter 16). Thus,
even discounting the prolonged survival findings
reviewed below, the tgo values for V. cholerae are not
greatly lower than those reported for coliforms and
may be similar to those reported for other bacterial
enteric pathogens. In a direct comparison of various
bacteria in sterile well water, McFeters and others
(1974) found the following t5 o values: shigellae, 22-27
hours; coliforms, 17 hours; salmonellae, 2-19 hours;

312

ENVIRONMENTAL BIOLOGY &EPIDEMIOLOGY: BACTERIA

Table 17-7. Survival of V. cholerae in sewage

Source
Altukhov
and others
(1975)P

Biotype and
initial concentration
per milliliter
Classical
(Ogawa)
lo0

Type of sample
Sewage of a bath
house (USSR;
BOD = 320 milligrams per liter)

37C

El Tor
2 x 10'

> 10 days

> 10 days

El Tor (Ogawa)
t0o
Daniel and
Lloyd (1980b)

Suruival6

Temperature

Strong sewage at
refugee camp
(Bangladesh)

22-25C

Non-Ol
(sewage isolate)
2 x 105

Concentration
fell by 1 log
in 6 hours and
remained steady
for further
42 hours
Concentration
rose to 4 x 106
in 6 hours and
remained steady
for a further
42 hours

Flu (1921)

Classical

Sewage in
septic tanks

Ambient
temperature
(Batavia)

2 days

Gerichter
and others
(1975)

El Tor

Sewage
(Jerusalem)

20-28C

Two phase
decline: t9 0 =
1.8 days for
first 5 logs
and tg, = 8
days subsequently.
V cliolerae not
detected after
24 daysc

Howard and
Lloyd (1979)

El Tor
106

Raw sludge
1 percent solids

25'C

t90 =

I day

5 percent solids

Kott and
Betzer
(1972)

El Tor
10

Diluted
sewage (Haifa:
BOD = 200 milligrams per liter)

Room temp.
(Israel)

Mukerjee.
Rudra and
Roy 11961)

Classical
2 x 106

Sewage (Calcutta)
Raw
Autoclaved
Filtered

Room temp.
(Calcutta)

El Tor
(clinical isolate)
2 x l0'

Raw
Autoclaved

2 days
max survival
14 days
t,, = 3 days
max survival > 14 days

1-5 days
4-24 days
2-7 days
2 days
9 days

313

VIBRIO CHOLERAE AND CHOLERA

Table 17-7 (continued)

Source

Biotype and
initial concentration
per milliliter

Type of sample

Temperature

Survivala

El Tor
(water isolate)
2 x 106

Raw
Autoclaved

2 days
10 days

Non-Ol
(clinical isolate)
2 x 106

Raw
Autoclaved

2 days
8 days

Non-Ol
(water isolate)
2 x 10'

Raw
Autoclaved

2 days
8 days

Ohwada
(1924); cited
by Pollitzer
(1959)

Classical

Sewage

Zaidenov
and others
(1976)

El tor (Ogawa)
104

10

Locomotive depot
wastewater
Domestic sewage
Dairy effluent
Oil and water
Diesel fuel and
water

4C
Room temp.
(Japan)
37C

12 days
4 days
1 day

18-24C

>39 days
3 days
14 days
> 14 months
> 14 months

Note: Older literature is reviewed by Pollitzer (1959).

a. Times given, for instance, as 6 days are durations at which viable organisms could not be detected. Times given as > 10 days indicate
that organisms were still viable at that time but that sampling was discontinued.
b. These experiments were discontinued after 10 days, at which time the concentration of classical V cholerae was 5 x 102 while that of El Tor
had risen to over 10' per milliliter. Data from the bath house suggested that V cholerae (El Tor, Ogawa I survived for at least 13 months in the
sewerage system (temperature 20-25C) despite repeated disinfection and no known external recontamination.
c. tgo: time for 90 percent reduction.

and V cholerae,7 hours. Pandit and others (1967)found

that V.cholerae (El Tor) survived2to 5timeslongerthan
E. coli, Pseudomonas spp., and Aerobacter spp. when
they were added to artificial well water and stored at
25 0 C.

of 9 x 105 per milliliter remained steady for 4 years for

both biotypes. Other results from these experiments
are given in table 17-4.
More remarkable are reports of prolonged survival
in raw sewage. Altukhov and others (1975) studied a
bath house in the USSR. V. cholerae (El Tor, Ogawa)
was isolated from 49 percent of samples of wastewater
from the bath house over a 13-month period. Repeated

Pollitzer (1959) cited several early studies that

reported prolonged survival of V. cholerae in various
waters. Examples are up to a year in sterilized spring or
well water, up to a year in sterilized river water, and
over 9 months in sterilized seawater.
Sayamov and Zaidenov (1978)'studied the survival
of classical and El Tor V. cholerae in mineral waters
from a spa at Matsesta (USSR). In raw mineral water,
survival did not exceed 22 days for either biotype. In
boiled mineral water at 20-24C, initial concentrations

attempts to disinfect the wastewater system had no

effect on V. cholerae isolation. There was no known
cholera infection in the community. V. cholerae was not
isolated from the incoming water supply, nor from large
numbers of samples of human feces, water, fish, and
frogs that were examined. Serological surveillance also
failed to detect evidence of V. cholerae infection. V.
choleraewas isolated from river water contaminated by
the discharge from the bath house. In laboratory
experiments, wastewater from the bath house

314

ENVIRONMENTAL BIOLOGY &EPIDEMIOLOGY: BACTERIA

(BOD 5 = 320 milligrams per liter, pH = 7.6) was

inoculated with an El Tor (Ogawa) strain previously
isolated from the bath house and with a reference strain
of classical V. cholerae (Ogawa), and stored at 37C.
The concentration of El Tor organisms was 105 per
milliliter at the start, rose to over 108 per milliliter after
3 days, and maintained this concentration up until 10
days when sampling was discontinued. The concentration of classical organisms was 103 per milliliter
at the start, rose to 105 after 3 days, and fell back to
5 x 102 after 10 days. The investigation failed to
discover how the bath house sewerage system became
infected, but it was clear that, once infection had taken
place, V. cholerae (El Tor, Ogawa) maintained itself in
the warm sewage (20-25C) and was remarkably
resistant to disinfection.
A very similar experience was reported by Zaidenov
and others (1976). A sewerage system serving a
locomotive depot and a housing estate was investigated. Wastewater from the locomotive depot (450
cubic meters per day) was rich in oil products and
passed through oil traps and a flotation chamber
before being mixed with domestic sewage (150-250
cubic meters per day). The mixed sewage was then
pumped to treatment fields. Because hot water was
used in the locomotive depot, the sewage was warm,
even in winter, and temperatures of 19-24C were
recorded throughout the year. The pH of the sewage

was 7.1 to 9.3. Over a 17-month period 1,454 samples of

sewage from various points in the system were
examined, and 17 percent were positive for V. cholerae
(El Tor, Ogawa). The wastewater from the locomotive
depot was far more frequently infected (18-42 percent)
than the domestic sewage (5 percent). The oil traps and
flotation chamber were most frequently infected. The
V. choleraestrain isolated was always the same and was
nontoxigenic. Fecal examination of 2,708 people in the
depot and the housing estate revealed only three
infections with non-O1 V. cholerae. When one oil trap
was isolated from the system, V. choleraewere shown to
survive in it for 36 days (the temperature in this oil trap
fell to 10C after isolation from the sewerage system).
In laboratory experiments, the El Tor strain isolated
from the locomotive depot was inoculated into various
wastewaters and stored at 18-24C. In mixtures of oil
plus water and diesel fuel plus water, survival was for
over 14 months, with an initial concentration of 10 per
milliliter. In domestic sewage, survival was less than 3
days; in locomotive depot wastewater, survival was
over 39 days; and in dairy effluent (included for
comparison), survival was less than 14 days. All
experiments were performed with initial inocula of 104
V. choleraeper milliliter. The source of infection of the
sewerage system was not discovered. Repeated
disinfection failed to clear V cRolerae from the network
until massive doses of chlorine (to achieve 10

Table 17-8. tg, values in hours for various types of V. cholerae in various waters and wastewaters
Classical 01

El Tor 01

Non-OI

Type of
water
enuironment

No.

Arith.
mean

Range

Arith.
mean

Range

No.

Arith.
mean

Range

Dechlorinated tap
water

22

3-48

49

2 163

ND

ND

ND

Well water

36

NA

13

116

5-264

ND

ND

ND

Surface water

18

0.16-36

10

53

1-230

8-8

Seawater

95

0.36-161

56

3-235

ND

ND

ND

Sewage

12

NA

66

8 240

8-8

Sterilized well
water, surface
water or sewage

34

3-65

59

32 168

39

31-50

No. Number of results.

Arith. mean Arithmetic mean.
ND No data.
NA Not applicable.

No.

VIBRIO CHOLERAE AND CHOLERA

milligrams per liter throughout the system) and

sulphuric acid (to lower sewage pH to 3-4) were added.
Following this, no V. cholerae were isolated for the next
12 months.
Further evidence of multiplication and prolonged
survival in some wastewater is provided by reports of
the multiplication of V. cholerae (El Tor, Inaba) in a
clinic septic tank in Japan (MMWR 1979) and the
multiplication of V. cholerae (non-O1) in a trickling
filter in Bangladesh (Daniel and Lloyd 1980b). These
occurrences, and their relationship to environmental
reservoirs of some atypical and non-Ol V cholerw.
await clarification.

Perhaps the greatest upset to traditional concepts of

cholera epidemiology and bacteriology has come from
the recent discoveries of V. cholerae and related
organisms occurring in surface waters not known to be
fecally contaminated or in areas where no human
infection has been recorded. V. cholerae, El Tor and
non-O1, were frequently isolated from wells, tanks, and
rivers in India in the 1930s and 1940s, but their close
relationship with classical V. cholerae 01, and their
potential pathogenicity, were not recognized at that
time (Read and Pandit 1941; Taylor and Ahuja 1938;
Venkatraman, Krishnaswami and Ramakrishnan
1941).
Colwell, Kaper and Joseph (1977) reported the
isolation of non-O1 V. cholerae from various parts of
Chesapeake Bay (USA). Subsequently, Kaper and
others (1979) described the ecology of non-O1 V.
cholerae in Chesaspeake Bay in some detail.
Concentrations were up to 7 per liter, and isolations
were only made at sites with salinities between 0.4 and
1.7 percent. There was no correlation between V.
cholerae counts and counts of total bacteria, coliforms,
fecal coliforms, or salmonellae. V. cholerae were not
especially associated with bottom sediment or oysters.
In a recent publication (Colwell and others 1980),
data on V. cholerae isolations from various brackish
and estuarine environments are summarized. V.
cholerae isolations in Chesapeake Bay were dependent on salinity and temperature, with the highest
recoveries (up to 46 per liter) being reported at
salinities of 0.3 to 1.7 percent and during the summer
when water temperatures were 28C. V. cholerae
isolations were not correlated with known fecal
contamination, nor with fecal coliform counts, thus
suggesting that V. cholerae "is an autochthonous
species in the estuarine ecosystem". Both non-O1 V.
cholerae serotypes and V. cholerae 01 (Inaba) have

315

been isolated from Chesapeake Bay. V. cholerae 01

(Inaba) has also been isolated from Louisiana salt
marshes. Some of the V. cholerae 01 and V. cholerae
non-O1 strains isolated from the Chesapeake Bay and
the Louisiana coast showed evidence of toxin
production. A marked association of V. cholerae non01 with zooplankton was found both in the
Chesapeake Bay and in surface water samples collected
in Bangladesh.
Bashford and others (1979) and West, Knowles and
Lee (1980) reported the isolation of up to several
hundred V. cholerae per milliliter from streams and
drainage ditches in Kent (England), including sites
where there was no known sewage contamination.
Isolations were more common during the summer.
Except for one occasion, all isolations have been of
non-O1 serotypes, and all have been nontoxigenic (J.
Lee, personal communication). Muller (1978, 1979)
isolated non-O1 V. cholerae from 33 percent of river
water samples in the Federal Republic of Germany, but
not from sewage treatment plant effluents. Isolations
were more numerous in summer.
V cholerae 01, atypical V cholerae 01 and non-Ol V
cholerae have been isolated variously from freshwater,
saline water, and wastewater in Australia, Bangladesh,
Brazil, England, Germany, Guam, Japan, the USA,
and the USSR (WHO Scientific Working Group 1980).
Most of these isolates have been found to be
nontoxigenic and nonpathogenic. They have been
found in areas where cholera cases or infections are not
known to occur (for example, Brazil, England, and the
USA) and in waters that are not thought to have
received any human fecal contamination (for example,
England and the USA). It is very probable that some of
these V. cholerae isolates are free-living aquatic
organisms. Whether they are in any way related to
human disease or to the epidemiology of cholera
remains to be determined.
The speculation concerning a possible environmental reservoir for atypical and non-O1 V. cholerae, and
possibly also for V. cholerae 01, has been increased by
findings on the affinity of these organisms for chitin.
Nalin and others (1979) found that about 70 percent of
V. cholerae 01 organisms, which were shaken for 6
hours with powdered crabshell in a 4.2 percent salt
solution at pH 6.2 and 20C. adsorbed to the chitin
particles. These adsorbed V. cholerae were then
somewhat resistant to an acid environment simulating
the stomach (pH) 1.6-1.8 for 13 minutes). V. cholerae
also multiplied (>4 log increase) when incubated
for 2 days at 37C in 4.2 percent salt solution
containing chitin. Other studies have shown that V.
cholerae 01 (classical and El Tor) and non-O1 can

316

ENVIRONMENTAL BIOLOGY &EPIDEMIOLOGY: BACTERIA

produce chitinase (Dastidar and Narayanaswami

1968) and that non-Ol V. cholerae, like V. parahlaemolvticus, can adsorb to, and multiply on, chitinous
fauna such as crab, shrimp, and zooplankton (Colwell,
Kaper and Joseph 1977; Kaneko and Colwell 1973,
1975, 1978; Kaper and others 1979; Nalin 1976;
Sochard and others 1979).
In sweat
Dodin and F&lix (1972) found that V. cholerae, El
Tor, was still viable after seven weeks at 28C in
human sweat and on gauze pads soaked in sweat and
stored in humid conditions. From one quantitative
experiment a tgo of 215 hours at 28C in sweat can be
computed. This is much longer than typical t90 values
at that temperature (table 17-8). Dodin and Felix

considered that these findings had considerable

relevance to the epidemiology of cholera in arid areas
of West Africa. Isaacson and Smit (1979) showed that
V. cholerae (El Tor, Inaba) multiplied, and could
survive for at least 120 hours, in pooled human sweat.
Multiplication of V. cholerae in sweat was believed to
have promoted the transmission of cholera among
South African gold miners undergoing heat acclimatization (Isaacson and others 1974). It is not known
whether V. cIIolera survives well in sweat on the skin.
On surlaces
V. cholerae survival on surfaces is usually limited
because of the sensitivity of the organism to
desiccation. Four studies on V. cholerae on various
household items are summarized in table 17-9.

Table 17-9. Survival of V. cholerae on surfaces

Source
Felsenfeld
(1965)

Biotype
Classical
and
El Tor

Type of surface
Absorbent materials
Cotton
Chopsticks
Paper
Shoes
Silk
Non-absorbent
materials
Aluminium foil
Coins
Tin cups
Plastic envelopes
China plates
Metal utensils

Temperature

Survtiala

28-30'C

5-7
2-3
2-3
2-3
3-5

days
days
days
days
days

28-30C

1 day
1 day
1 day
1-2 days
1-2 days
1-2 days

Gohar and
Makkawi
(1948)

Classical

Linen
Wool
Leather
Paper and rubber
Coins

Room temp.
(Egypt)

6 days
5 days
3 days
10 hours
6 hours

Pesigan,
Plantilla
and Rolda
(1967)

El Tor

Frying pan
China plates
Pestle and mortar
Drinking glass
Metal utensils
Kitchen knife
Wooden chopping
block

30-32C

4 hours
4 hours
4 hours
4 hours
24 hours
48 hours
24 hours

Shousha (1948)

Classical

Cotton and cloth

Bank note
Postage stamp
Coin

Room temp.
(Egypt)

4
3
2
I

Note: Older literature is reviewed by Pollitzer (1959).

a. Times given are those at which viable organisms could no longer be detected.

days
days
days
day

VIBRIO CHOLER4E AND CHOLERA

The longer persistence on absorbent materials,

especially cotton, is interesting and suggests that
clothing (especially clothing soaked in sweat) may act
as a temporary habitat for V. cholerae. It is also
noteworthy that survival times are markedly shorter
than those reported for other enteric bacteria-for
instance, Shigella (chapter 16)-on similar surfaces.

317

likely that some foods can and do act as a primary

vehicle for spreading cholera, especially within the
household or at feasts and markets.

Inactivation by Sewage Treatment

Processes
In soil

There is very little information on the fate of V

cholerae in sewage treatment plants partly because, as
ih hlr
rdc
metoe abve motpol
Experiments in Israel (Gerichter and others 1975)
mentioned above, most people with cholera produce
found that V. cholerae (El Tor )in soil survived for up to
no sewage; therefore V. cholerae is only very rarely
y
y
y
g
4 days when the soil was allowed to dry slowly, but for
up t dy h howr afound
in sewage, and even then in low concentrations.
Flu (1921) studied seeded V. cholerae in septic tanks
wupnton10tdaysmwenathe sowas regulal c
remois
in Batavia (now Jakarta; Indonesia). A total of five
with uncontaminated sewage (initial concentrations
setcanswrsudd,ndiolyneasV
were 10 per gram of soil, and the storage temperature
was 20-28C). Nalin and others (1980) reported
septic tanks were studied, and in only one was V.
cholerae detected in the effluent. Early studies reviewed
wasurviv
al
dayswhendV.
inorover6
hotherse(1980) rportd
w
survival for over 6 days when V. cholerae (El Tor) was
inoculated into sterile potting soil and stored at 26C.
by Kabler (1959) reported a 98 percent reduction of V.
cholerae in an activated sludge plant.
inocuatedinto
terie poting oil ad streda 6
sludge ant.
Kott an Beted
In the same experiments it was found that common
Kott and Betzer (1972) studied a 70-liter model
earthworms (Luimbricus terrestris)ingested V cholerae
in sol ad swaste stabilization pond with a retention time of 5
days. The pond was fed with diluted sewage
in soil and subsequently died. V. choleraemultiplied in
nigams
.wihV
per lite siked
days. T 20
the earth worms and were isolated at concentrations
(BOD5 = 200 millilgrams per liter) spiked with V.
up
to
pe
.0.illtro
wr
ooeae
up to 10~ per milllihter of worm homogenate.
cholerae (El Tor). Influent coliform and V cholerae

On food and crops

In looking at the potential of food for transmitting
cholera, it is important to make the distinction between
food that acts as a primary vehicle for cholera,
becoming infected through direct contact with the
stools of a case or carrier, and food that acts as a
secondary vehicle of spread, becoming contaminated
by polluted water. Most documented occurrences of
foodborne cholera are of the second kind, and the most
numerous of these incidents are those involving fish
and shellfish. Alternatively, food can act as a secondary
vehicle of cholera through the use of polluted water to
irrigate or freshen vegetables.
The evidence for foods acting as the primary vehicles
for cholera is very limited. This is to be expected
because few studies have examined the domestic
environment in a cholera area during an outbreak and
carried out a systematic investigation of food for V.
cholerae. Table 17-10 summarizes some literature on
the survival of V. cholerae on food. It is clear that
survival times of several days are commonly achieved,
even at around 30C. Survival is longest in moist,
nonacidic, and sterile (that is, cooked) foods. Only two
studies (Felsenfeld 1965; Neogy 1965) directly compared the survival of the classical and El Tor biotypes,
and both found that El Tor survived for longer. It seems

concentrations
were
3 x 106'8 x 108
and
1 x 103-8 x 103 per 100 milliliters, respectively.
Effluent coliform and V. cholerae concentrations were
8 x 104-4 x 107 and 0-2 per 100 milliliters respectively. The addition of 8 milligrams per liter of
chlorine to the waste stabilization pond effluent
eliminated all remaining V. cholerae.
Daniel and Lloyd (1980a) studied two Oxfam
Sanitation Units in refugee camps near Dacca
(Bangladesh). These units treated very strong sewage
(17,000 and 7,400 milligrams of suspended solids per
liter) in two unbaffled, flexible tanks connected in
series. Each tank had a volume of 18 cubic meters, and
the flow of sewage was 2.5 to 3 cubic meters per day.
Thus, the total mean retention times were 12-15 days.
The geometric mean inflowing concentrations of non01 V cholerae at the two camps were 2.6 x 103 and
1.6 x 102 per 100 milliliters, respectively. The geometric mean effluent concentrations were 6.5 and 5.3 per
100 milliliters. Thus, overall removal rates at the two
camps were 99.8 and 96.4 percent, respectively. These
removal rates give tgo values of 106 and 257 hours,
respectively, which are longer than those reported in
table 17-7, especially if the warm ambient temperature
is taken into account. This suggests either shortcircuiting in the tanks, which is quite probable, or non0 1 V. cholerae multiplication in the warm sewage in the
tanks.

318

ENVIRONMENTAL BIOLOGY &EPIDEMIOLOGY: BACTERIA

Table 17-10. Survival of V. cholerae on food and crops

Source

Biotype

Type offood

Temperature

SurviualI

.4. Meat
Cheng (1963)

El Tor

Beef

Day 1: 22C
Thereafter: 3-4C

5 days

Felsenfeld
(1965)

El Tor and
classical

Raw beef

2-4C
28-30C
2-4C
28-30C

5-7
1-2
1-2
3-7

Sausages
(surface and inside)

2-4C
28-30C

1 day
1 day

Raw meat

5-10C
30-32C

4-9 days
2-4 days

Cooked meat

5-I0'C
30-32C

3-5 days
2 5 days

Cooked beef

Pesigan,
Plantilla
and Rolda
(1967)

El Tor

days
days
weeks
days

B. Fish
Cheng (1963)

El Tor

Lice-eye fish
Sliced sword-fish

Day 1: 21.5C
Thereafter: 4C

16 days
10 days

Felsenfeld
(1965)

El Tor and
classical

Shrimp

2-4C
28-30C

1-3 days
1-2 days

2 4C
28-30C

1-2 weeks
2-4 days

Dried

2-4C
28-30C

3-5 days
1-2 days

Salted

2-4C
28-30C

1-2 days
I day

Cooked

2-4C
28-30C

2-7 days
1-6 days

5-10C
30-32C

4-9 days
2-4 days

Catfish
Raw

Pesigan,
Plantilla
and Rolda (1967)

El Tor

Various fish and

shellfish

C. Vegetables andfruit
Cheng (1963)

El Tor

Horseradish
Cucumber
Tomato
Orange

Day 1: 22C
Thereafter: 3-4C

21 days
23 days
16 days
14 days

El Shawi and
Thawaini
(1967)

El Tor

Date
Melon

Room temp.
(Iraq)

3 days
2 days

Felsenfeld
(1965)

El Tor and
classical

A comprehensive
survey of a wide range
of cooked and
uncooked fruits and
vegetables

2-4C
28-30C

Up to 4 weeks
Up to 7 days (except inside melon,
which was 2 weeks); survival
was especially long on cabbage,
cucumber, eggplant, melon,
okra, peas, and potatoes.

319

VIBRIO CHOLERAE AND CHOLERA

Table 17-10 (continued)

Source
Gerichter
and others
(1975)

Biotype
El Tor

Type offood
Parsley
Tomato and carrot
Cucumber, pepper,
and okra
Lettuce

Temperature
20-26'C

Survivala
1 day
1.5 days
1-2 days
2-3 days
Mean death rates for all the above
were 4-6 log units per day

Parsley
Wet
Dry

20-28C

2 days
1 day

Lettuce
Group of leaves
Single leaf

18-26C

68 hours
44 hours

22-30C

4 hours

Tomato
in sunlight
Parsley
Lettuce

4C

2 days
4 days

Gohar and
Makkawi (1948)

Classical

Date
Vegetables

Room temp.
(Egypt)

4 days
6 days

Neogy
(1965)

El Tor and
classical

Papaya
Cucumber
Pineapple
Boiled rice
soaked overnight

Room temp.
(India)

1 day
>1 day
15 minutes

Cooked fruit and

vegetables

5-1OIC
30-32C
5-10C
30-32C
5-10C
30-32C

3-5 days
2-5 days
2-3 days
1 day
6-9 days
2-5 days

20-25C

1 hour

Pesigan,
Plantilla
and Rolda
(1967)

El Tor

Prescott
and
Bhattacharjee
(1969)

El Tor

Fresh fruit
Fresh vegetables
Lime, lemon, and
date
Orange, grape,
fig, raisin, and
tomato

1 hour

1 day

Banana, guava,
papaya, onion, eggplant,
pea, celery, green bean.
bean sprout, and rice.
Okra, lima bean,
pumpkin, and
potato
Shousha
(1948)

Classical

Onion and date

Garlic, rice,
lentil, and grape
Orange and lemon

2-5 days
6-8 days
Room temp.
(Egypt)

4 days
3 days
7 hours

320

ENVIRONMENTAL BIOLOGY & EPIDEMIOLOGY: BACTERIA

Table 17-10 (continued)

Source

Biotype

Type of food

Temperature

Survivala

D. Milk and Milk products

Felsenfeld
(1965)

El Tor and
classical

Butter,
unsalted
Cheese

2-4C
28-30C
2-4'C
28-30C

1-2 weeks
1 week
2-3 weeks
I week

Custard

2-4 DC
28-30C

3-4 weeks
1-2 weeks

Ice cream

2-40 C
28-30C

3-4 weeks
5-7 days

Milk

2-4C
28-30'C

3-4 weeks
1-3 weeks

Lema, Ogawa
and Mhalu
(1979)

El Tor

Milk

4C
30C

3 weeks
4 days

Neogy
(1965)

El Tor and
classical

Milk desserts

Room temp.
(India)

I day

Pesigan,
Pantilla
and Rolda
(1967)

El Tor

Milk, ice cream,

and butter

5-10'C
30-32C

I week->2 weeks
5-14 days

Prescott and
Bhattacharjee

El Tor

Milk desserts

20-25'C

1-2 days

Shousha
(1948)

Classicat

Milk
Sour milk
Butter
Cheese

4C
Room temp.
[_
rr)

> 2 days
2 hours
>2 days
7 hours

El Shawi
and Thewaini
(1967)

El Tor

Barley and
wheat

Room temp.
(Iraq)

2 days

Felsenfeld
(1965)

El Tor and
classical

A comprehensive
survey of a
wide range of
cooked and
uncooked foods

2-4C
28-30C

Up to 4 weeks
Not more than
7 days,
except for
coconut cream (10 days),
coconut dishes (3 weeks).
and noodles (2 weeks)

Gohar and
Makkawi (1948)

Classical

Honey and
treacle

Room temp.
Fl -. I

3 hours

Neogy (1965)

El Tor and
classical

Sweet and sour

curd
F . n and
sandesh

Room temp.
(India)

5 minutes

Cooked noodles.
rice cake, and
jam

5-10C
30-32CC

(1969)

E. Other foods

Pesigan.
Plantilla
and Rolda
(1967)

El Tor

1 day

3-5 days
2-5 days

321

VIBRIO CHOLERAE AND CHOLERA

Table 17-10 (continued)

Source

Biotype

Type offood

Temperature

Survivala

Prescott and
Bhattacharjee
(1969)

El Tor

Wheat and nuts

Spices

2025aC
20-25C

3 days
1-5 days

Shousha
(1948)

Classical

Sugar
Bread
Honey

Room temp.
(Egypt)

4 days
3 days
2 days

F Beverages
El Shawi
and Thewaini
(1967)

El Tor

Soft drinks

Room temp.
(Iraq)

1 day

Felsenfeld
(1965)

El Tor and
classical

Beer, carbonated
water,
carbonated soft drinks,
ime and whisky
Cocoa

2-4C
28-30C

I day
I day

2-4C
28-30'C

1-2 weeks
3-5 days

Coffee

2-4C
28-30C

1-2 days
1 day

Lema, Ogwa
and Mhalu
(1979)

El Tor

Ice cubes

2-4C

4-5 weeks

Lemonade

2-4C
28-30C

2-3 weeks
5-7 days

Tea

2 4C
28-30C

1 week
2-3 days

Coconut fluid

4'C
3OcC
4C

4 days
2 days
I hour
(except for
mnbege, in which
survival was
2 days)

30C

I hour

Beer, gin, and

traditional
alcoholic
beverages chibuku
(maize and beans)
and mbege
(bananas and millet)

Pesigan,
Pantilla
and Rolda
(1967)

El Tor

Coca cola

5-1OC
30-32C

2 days
4 hours

Prescott and
Bhattacharjee
(1969)

El Tor

Coca cola
Rosewater
Ground coffee
Tea leaves

20-25WC

1 day
2 days
I hour
I day

Note: Older literature is reviewed by Pollitzer (1959).

a. Times given, for instance, as 2 days are durations at which viable organisms could no longer be detected. Times given as > 2 days indicate
that organisms were still viable at that time but that sampling was discontinued.

322

ENVIRONMENTAL BIOLOGY &EPIDEMIOLOGY: BACTERIA

ZONE

45-

t-45

SAFETY

40

- 40

35-

-35

~ ~~

x x~

x0

fi20 -

15-

,15

~=xXx-

20

_15

X 100h destruction of Vibrio cholerae

10

e*less than 100%/c

destrvction of Vibrio cholerae

5-

-10

*
1

wxOx

10

100

iday

1000

Iweek
TIME

-5
lmonth

10000

lyear

(HOURS)

Figure 17-3. The influence of time and temperatureon V. cholerae. The points plotted are the results of experiments
done under widely differing conditions. The line drawn represents a conservative upper boundary for death

Daniel and Lloyd (1980b) reported that small

trickling filters were installed to treat further the
effluents from these Oxfam Sanitation Units. Influent
concentrations (effluents from the second tank of the
main unit (non-O1 V. cholerae were 3-9 per 100
milliliters, while effluents from the trickling filters
contaied
pr3-,400
100milliiters.The athors
contained 3-2,400 per 100 milliliters. The authors
concluded that non-O1 V. cholerae was multiplying in
ponded sewage in the trickling filters.

Literature Cited
Abel, R. and Claussen, R. (1895). Untersuchungen uiber die
lebensdauer der cholera-vibrionen in fakalien. Zentralblatt
fur Bakteriologie, und Parasitenkunde, I, 17, 77-81 and
118-130.
Altukhov, A.A., Ivanov, S.I., Semiotrochevl, V.L., Bulegenov,
E. I., Lebedev, K. K., Filimonova, L. V., Zakharov, N. I.
and Rybak, E. t. (1975). Prolonged detection of El Tor
cholera vibrio in sewage of a bath-house. Zhurnal
Mikrobiologii, Epidemiologii i Immunobiologii, 2, 41-44.
Azurin, J. C. and Alvero, M. (1974). Field evaluation of

Inactivation by Night Soil and

environmental

Sludge Treatment Processes

Bulletin of the World Health Organization, 51, 19-26.

sanitation measures against cholera.

Azurin, J. C., Kobari, K., Barua, D., Alvero, M., Gomez, C. Z.,
No reports of V.cholerae reduction during night soil
Dizon, J. J., Nakano, E-I., Suplido, R. and Ledesma, L.
or sludge treatment were located. The data given in
(1967). A long-term carrier of cholera: Cholera Dolores.
table 17-6 suggest that V. cholerae will be eliminated by
Bulletin of the World Health Organization, 37, 745-749.
,,hn e icBart, K. J., Khan, M. and Mosley, W. H. (1970). Isolation of
anyoproestha
hing
as
awaretenimatio
Time-ofappreiaby
Vibrio cholerae from nightsoil during epidemics of classical
and El Tor cholera in East Pakistan. Bulletin of the World
combinations lethal to V. cholerae are given in figure
Health Organization, 43, 421-429.
17-3. It appears that V. cholerae will be eliminated by
Bashford, D. J., Donovan, T. J., Furniss, A. L. and Lee, J. V.
almost any sludge digestion, composting, or storage
(1979). Vibrio cholerae in Kent. Lancet, 1, 436-437.
process and will certainly be removed far more readily
Blake, P. A., Rosenberg, M. L., Florencia, J., Costa, J. B.,
than E. coli and other fecal indicator bacteria (chapter
Quintino, L. D. P. and Gangarosa, E. J. (1977). Cholera in
13).
Portugal, 1974. II. Transmission by bottled mineral water.

VIBRIO CHOLERAE AND CHOLERA

American Journal of Epidemiology, 105, 344-348.

Briscoe, J. (1978). The role of water supply in improving
health in poor countries (with special reference to Bangla
Desh). American Journial of Clinical Nutrition, 31,
2100-2113.
Cash, R. A., Music, S. I., Libonati,J. P.,Snyder,M.J.,Wenzel,
R. P. and Hornick, R. B. (1974). Response of men to
infection with Vibrio cholerae. I. Clinical, serologic and
bacteriologic responses to a known inoculum. Journal of
Infectious Diseases, 129, 45-52.
Cheng, C. T. (1963). Survival of El Tor vibrio outside the
human body. Nagasaki Igakkai Zassi, 38, 870-876.
Colwell, R. R., Kaper, J. and Joseph, S. W. (1977). Vibrio
cholerae, Vibrio parahaemolyticus, and other vibrios:
occurrence and distribution in Chesapeake Bay. Science,
198, 394-396.
Colwell, R. R., Kaper, J., Seidler, R., Voll, M. J., MeNicol, L.
A. Garges, S., Lockman, H., Maneval, D. and Remmers, E.,
Joseph, S. W., Bradford. H., Roberts, N., Huq, I. and Huq,
A. (1980). Isolation of 01 and non-Ol Vibrio cholerae from
estuaries and brackish water environments. In Proceedings
of the 15th Joint Conference on Cholera, US-Japan
Cooperative Medical Science Program, pp. 44-51.
Bethesda, Md.: National Institutes of health.
Cvjetanovic, B. (1979). Sanitation versus immunization in
control of enteric and diarrhoeal diseases. Progress in
Water Technology, 11, 81-87.
Cvjetanovic, B., Grab, B. and Uemura, K. (1978). Dynamnics of
Acute BacterialDiseases, EpidemiologicalModels and their
Application in Public Health. Geneva: World Health
Organization.
Daniel, R. R. and Lloyd, B. J. (19 80a). Microbiological
studies on two Oxfam Sanitation Units operating in
Bengali refugee camps. Water Research, 14, 1567-1571.
(1980b). A note on the fate of El Tor cholera and other
vibrios in percolating filters. Journal of Applied
Bacteriology, 48, 207-210.
Dastidar, S. G. and Narayanaswami, A. (1968). The
occurrence ofchitinasein vibrios.IndianJournalofMedical
Research, 56, 654-658.
Dizon,J., Fukumi, H., Barua, D., Valera,J.,Jayme,F., Gomez,
F., Yamamoto, S-I., Wake, A., Gomez, C. Z., Takahira. Y.,
Paraan, A., Rolda, L., Alvero, M., Abou-Gareeb, A. H.,
Kobari, K. and Azurin, J. C. (1967). Studies on cholera
carriers. Bulletin of the World Health Organization, 37,
737-743.
Dodin, A. and Felix, H. (1972). Du role de la sueur dans
l'epidemiologie du cholera en pays sec. Bulletin de
l'Academie Nationale de Medicine, 156, 845-852.
El-Shawi, N. N. and Thewaini, A. J. (1967). El Tor vibrios
isolated in Iraq and their survival on some foods and
beverages. WHO/Cholera Information/67.9 Geneva:
World Health Organization, Unpublished document.
Feachem, R. G. A. (1976). Is cholera primarily water-borne?
Lancet, 2, 957.
(1981). Environmental aspects of cholera epidemiology. I. A review of selected topics reports of endemic and

323

epidemic situations during 1961-1980. Tropical Diseases

Bulletin. 78, 675-698.
(1982). Environmental aspects of cholera epidemiology. III. Transmission and control. Tropical Diseases
Bulletin, 79, 1-47.
Felsenfeld, 0. (1965). Notes on food, beverages and fomites
contaminated with Vibrio cliolerae. Bulletin of the World
Health Organization, 33, 725-734.
(1974). The survival of cholera vibrios. In Cholera,
eds. Barua, D. and Burrows, W., pp. 359-366. Philadelphia:
W. B. Saunders.
Flu, P. C. (1921). Investigations of the duration of life of
cholera vibriones and typhoid bacteria in septic tanks at
Batavia.
Mededeelingen
van
der
Burgerlijken
Geneeskundigen Dienst in Nederlandschindie, 3, 289-297.
Forbes, G. I. Lockhart, J. D. F. and Bowman, R. K. (1967).
Cholera and nightsoil infection in Hong Kong, 1966.
Bulletin of the World Health Organization,36, 367-373.
Gangarosa, E. J. and Mosley, W. H. (1974). Epidemiology
and surveillance of cholera. In Cholera,eds. Barua, D. and
Burrows, W., pp. 381-403. Philadelphia: W. B. Saunders.
Gerichter, C. B., Sechter, I. Gavish, A. and Cahan, D. (1975).
Viability of Vibrio cholerae biotype El Tor and of cholera
phage on vegetables. Israel Journalof Medical Science, 11,
889-895.
Gildemeister, E. and Baerthlein, K. (1915). Beitrag zur
cholerafrage. Mainchener Medizinische Wochenschrift, 62,
705-708.
Gohar, M. A. and Makkawi, M. (1948). Cholera in Egypt:
Laboratory diagnosis and protective inoculation. Journal
of Tropical Medicine anid Hygiene, 51, 95-99.
Greenough, W. B. (1979). Vibrio cholerae. In Principlesand
Practice of Infectious Diseases, eds. Mandell, G. L.,
Douglas, R. G. and Bennett, J. E., pp. 1672-1687. New
York: John Wiley.
Grieg, E. D. W. (1914). On the vitality of the cholera vibrio
outside the human body. Indian Journal of Medical
Research, 1, 481-504.
Gunn, R. A., Kimball, A. M., Pollard, R. A., Feeley, J. C. and
Feldman, R. A. (1979). Bottle feeding as a risk factor for
cholera in infants. Lancet, 2, 730-732.
Hornick, R. B., Music, S. 1., Wenzel, R., Cash, R., Libonati, J.
P., Snyder. M. J. and Woodward, T. E. (1971). The Broad
Street pump revisited: response of volunteers to ingested
cholera vibrios. Bulletin of the New York Academy of
Medicine. 47, 1181-1191.
Howard, J. and Lloyd, B. (1979). Sanitation and disease in
Bangladesh urban slums and refugee camps. Progress in
Water Technology, 11, 191-200.
Isaacson, M., Clarke, K. R., Ellacombe, G. H.. Smit.
W. A., Smit, P., Koornhof, H. J., Smith, L. S. and Kriel, L. J.
(1974). The recent cholera outbreak in the South African
gold mining industry. South African Medical Journal, 49,
2557-2560.
Issacson, M. and Smit, P. (1979). The survival and
transmission of V. cholerae in an artificial tropical
environment. Progress in Water Technology, 11, 89-96.

324

ENVIRONMENTAL BIOLOGY & EPIDEMIOLOGY: BACTERIA

Jamieson. W., Madri, P. and Claus, G. 11976). Survival of

certain pathogenic microorganisms in sea water.
Hydrobiologia, 50, 117-121.
Kabler, P. (1959). Removal of pathogenic micro-organisms
by sewage treatment processes. Sewage and Industrial
Wastes, 31, 1373-1382.
Kaneko, T. and Colwell, R. R. (1973). Ecology of Vibrio
parahaemnolyticus in Chesapeake Bay. Journal of
Bacteriology, 113, 24-32.
(1975). Adsorption of l'ibrio parahaemnolyticus onto
chitin and copepods. Applied Microbiology. 29, 269-274.
(1978). The annual cycle of Vibrio paralhaemiolvticus
in Chesapeake Bay. Microbial Ecology. 4, 135-155.
Kaper, J.. Lockman, H.. Colwell. R. R. and Joseph, S. W.
(1979). Ecology, serology, and enterotoxin production of
Vibrio cliolerae in Chesapeake Bay. .4pplied aniid
Environmental Microbiology, 37, 91-103.
Khan, M. and Shahidullah. M. (1980). Cholera due to the El
Tor biotype equals the classical biotype in severity and
attack rates. Journalof' Tropical Medicine and Hygiene. 83,
35-39.
Khan, S. and Agarwal. M. N. (1929). On the duration of the
life of vibrios in the Ganges and Jumna river water. Inldiani
Journial of'Medical Research, 16, 993-1008.
Koch, R. (1884). An address on cholera and its bacillus.

British Medical Journal, August 30, 403-407 and 453-459.

Konchady. D. Prescott, L. M., Datta, A. and De, S. P. (1969).
Survival of Vibrio clholerae in some of the waters of
Calcutta. Inidian Journal of'Medical Researclh, 57, 60-65.
Kott, Y. and Betzer, N. (1972). The fate of Vibrio cholerae (El
Tor) in oxidation pond effluents. Israel Journlal ofMedical
Science. 8, 1912-1916
Lahiri, M. N., Das, P.C. and Malik, K. S. (1939). The viability
of Vibrio chlolerae in natural waters. Indiat. Medical
Gazette. 74, 742-744.
Lema, O., Ogwa, M. and Mhalu, F. S. (1979). Survival of El
Tor cholera vibrio in local water sources and beverages in
Tanzania. East African Medical Journal, 56, 504-508.
Levine, R. J. and Nalin, D. R. (1976). Cholera is primarily
waterborne in Bangladesh. Lancet. 2. 1305.
McFeters, G. A.. Bissonnette. G. K.. Jezeski, J. J., Thomson,
C. A. and Stuart, D. G. (1974). Comparative survival of
indicator bacteria and enteric pathogens in well water.
Applied Microbiology. 27, 823-829.
Mhalu, F. S.. Mmari, P. W. and Ijumba. J. (1979) Rapid
emergence of El Tor Vibrio cliolerae resistant to
antimicrobial agents during first six months of fourth
cholera epidemic in Tanzania. Lancet, 1, 345-347.
MMWR (1979). Cholera surveillance-Japan. Miorbidirv and
It./i. ;,,, Weekly Report. 29, 98-99.
Mukerjee, S., Rudra, B. C. and Roy, U. K. G. (1961).
Observations on cholera endemicity in Calcutta and
survival of Vibrio cholerae in the water sources. Annials of
Biochemistry and Experiniental Medicine, 11, 31-40.
Muller, H. E. (1978). Occurrence and ecology of NAG vibrios
in surface waters. Zentralblatt foir Bakteriologie.
Parasitenikuinde, Inifektions krankheiten und Hygiene, 1. B.
167, 272-284.

(1979). NAG-vibrionen. choleraahnliche erreger in

unseren gewassern. Forum Stidte-HlYgiene, 30, 211-214.
Nalin, D. R. (1976). Cholera, copepods and chitinase, Lanicet,
2, 958.
Nalin, D. R., Dava, V., Reid. A., Levine, M. M. and Cisneros.
L. (1979). Adsorption and growth of Vibrio cliolerae on
chitin. Infection and Immutnity. 25, 768-770.
Nalin, D. R., Robbins-Browne, R., Levine, M. M.. Daya, V.
and Noble, P. (1980). Multiplication of Vibrio cholerae in
earthworms. In Curreiti Ci, .. l .
and Infectious
Disease: Proceedingsof'the 11th InternationalCongress of'
Chemvotherapy and the 19th Ilnterscienzce Conference on
Anti-Microbial Agents and Chemotherapy, eds. Nelson, J.

D. and Grassi, C., pp. 936-937. Washington. D.C.:

American Society of Microbiology.
Neogy, K. N.. (1965). Viability of V.clolerae and V.cholerae
El Tor in food and water. Bulletini of'thle Calcutta School of
Tropical Medicine, 13, 10-11.
Ohwada, S. (1924). Destination of cholera vibrios in the
sewage watcr of Tokyo city. Journalof the Kejo Medical
Society. 3, 11-12.
Pandit, C. G., Pal, S. C., Murti, G. V., Misra, B. S., Murty, D.
K. and Shrivastav, J. B. (1967). Survival of V. cliolerae
biotype El Tor in well water. Bulletin of the World Health
Organiiation, 37, 681-685.

Pesigan, T. P., Plantilla, J. and Rolda, M. (1967). Applied

studies on the viability of El Tor vibrios. Bulletini oj'tlhe
World Health Organization, 37. 779-786.
Pollitzer, R. (1959). Cholera. Monograph 43. Geneva: World
Health Organization.
Prescott, L. M. and Bhattacharjee, N. K. (1969) Viability of
El Tor vibrios in common foodstuffs found in an endemic
cholera area. Bulletin of thle World Health Organzization, 40,
980-982.
Read, W. D. B.. Gurkirpal Singh. J., Seal, S. C. and Bose. S.
(1939). Growth and survival of V. chlolerae with special
reference to growth and survival in water. IndianiJournalof
Medical Researchl, 27, 1-40.
Read, W. D. B. and Pandit, S. R. (1941). Distribution of V.
cholerae and El Tor type strains in certain rural areas in
India. Indian Journlal of'Medical Researclh, 29, 403-415.
Sack, G. H.. Pierce, N. F., Hennessey. K. N., Mitra. R. C.,
Sack, R. B. and Mazumder, D. N. G. (1972). Gastric acidity
in cholera and non cholera diarrhoea. Bulletin of the World
Health Organization, 47. 31-36.
Sanyal. S. C.. Singh. S. J., Tiwari, 1.C., Sen, P. C.. Marwah, S.
M., Hazarika, U. R., Hardas Singh, Shimada, T. and
Sakazaki, R. (1974). Role of household animals in
maintenance of cholera infection in a community. Journal
of Infectiouis Diseases, 130, 575-579.
Sayamov, R. M. and Zaidenov, A. M. (1978). Survival and
properties of cholera vibrios cultivated in mineral water.
Zhurnal Mikrobiologii, Epidemiologii i 1
.i.,
,,, 11,
66-70.
Shoda, T., Koreyeda, T. and Otomo, T. (1934). The viability
of cholera vibrios in the human excreta. .Journal of the
Public Health Association of Japan, 10, 1-9.
Shousha. A. T. (1948). Cholera epidemic in Egypt (1947): a

I IBRIO

CHOLERAE AND CHOLERA

preliminary report. Bulletin ol the llorld Healthi

Organization, 1, 353--381.
Shrewsbury, J. F. D. and Barson, G. J. (1957). On the
absolute viability of certain pathogenic bacteria in a
synthetic well-water. Journlal ol Pathology and
Bacteriology, 74, 215-220.
Sinha, R.. Deb, B. C.. De, S. P., Abou-Gareeb, A. H. and
Shrivastava, D. L. (1967). Cholera carrier studies in
Calcutta in 1966-67. Bulletin of the World Health
OrganiZation, 37, 89--100.
Smith, H. L., Freter, R. and Sweeney, F. J. (1961).
Enumeration of cholera vibrios in fecal samples. Journalof
InJectious Diseases. 109, 31-34.
Sochard, M. R., Wilson, D. F., Austin, B. and Colwell. R. R.

(1979). Bacteria associated with the surface and gut of

marine
copepods.
Applied
and
Environmental
Microbiology, 37, 750-759.

Sommer, A. and Mosley. W. H. (1973). Ineffectiveness of

cholera vaccination as an epidemic control measure.
Lancet, 1, 1232-1235.
Taylor, J. and Ahuja, M. L. (1938). Incidence and characters
of vibrios in waters in northern India. Indian Journal oJ
Medical Research, 26, 1-32.
Threlfall, E. J., Rowe, B. and Huq, 1.(1980). Plasmid-encoded
multiple antibiotic resistance in Vibrio cholerae El Tor
from Bangladesh. Lancet, 1, 1247-1248.
Towner, K. J., Pearson, N. J., Mhalu, F. S. and O'Grady, F.
(1980). Resistance to antimicrobial agents of Vibrio
cholerae El Tor strains isolated during the fourth cholera
epidemic in the United Republic of Tanzania. Bulletin o.
the World Health Organiization, 58, 747 -751.

325

Towner, K. J., Pearson, N. J. and O'Grady, F. (1979).

Resistant Vibrio cholerae El Tor in Tanzania. Lancer, 2,
147-148.
Van de Linde, P. A. M. and Forbes. G. l. (I 9651. Observations
on the spread of cholera in Hong Kong, 1961--1963.
Bulletin of tile World Health Organlizationi,32. 515-530.
Venkatraman. K. V., Krishnaswami, A. K. and
Ramakrishnan,C. S. (1941). Occurrence of vibrio El Tor in
natural sources of water in the absence of cholera. Indian
Journial of Medical Researchi, 29, 419-424.
West, P. A.. Knowles, C. J. and Lee, J. V. (1980). Ecology of
Vibrio species, including Vibrio cholerae,in waters of Kent,
United Kingdom. Society for General Microbiology
Quarterly, 7, 80.
WHO Scientific Working Group (1980). Cholera and other
vibrio-associated diarrhoeas. Bulletinz of the World Health
Organization, 58, 353-374.

Yasukawa, Y. (1933). Experiments on sea water and Vib7io

cholerae. JapaneseJournlal of Experimnental Medicinie, 11,
119-127.
Zaidenov. A. M., Sayamov, R. M., Maloletkov, 1. S.,
Lazorenko, N. F., Bichul, K. G., Milyutin, V. N., Titenko,
M. T.. Popov. G. M., Kotov, V. K., Galtseva, G. V..
Goldberg, A. M.. Kiseleva. V. l., Stepanets, V. 1.,
Seliverstov, V. I., Goryachkina. T. Ya., Reznikov. L. G..
Lukash, V. O., Arendarskava, L. S. and Azykovskaya, 1.S.
(1976). Prolonged survival of El Tor cholera vibrio in
naturally infected sewage. Zhu1rnal Mikrobiologii.
Epidemiologii i Iinniunobiologii,12, 61-69.