Transcript Salmonella Praveen Rao, Sophia W. Riccardi, Danielle Birrer Seminar in Nucleic Acids-Spring 2004 Prof.

Salmonella
Praveen Rao, Sophia W. Riccardi, Danielle Birrer
Seminar in Nucleic Acids-Spring 2004
Prof. Zubay
Salmonella
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Overview
History and Epidemiology
Molecular Biology
Clinical
Weaponization
Overview
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Salmonella is a rodshaped, gramnegative, facultative
anaerobe in the
family
Enterobacteriaceae
Salmonella Taxonomy
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The genus Salmonella is divided into two species, S. enterica and S. bongori
(CDC).
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Over 2000 strains are grouped into S. enterica. This species is further
divided into six subgroups based on host range specificity, which also
involves immunoreactivity of three surface antigens, O, H and Vi.
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All strains that are pathogenic to humans are in species S. enterica, subgroup
1 (also called enterica).
For example, the correct taxonomic name for the organism that causes
typhoid fever is Salmonella enterica ssp. enterica, serovar typhi. The
simplified version: Salmonella typhi.
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Taxonomy has been revised several times, due to the degree of DNA
similarity between genomes.
For example, In the U.S., another legitimate species name for enterica is
choleraesuis.
Other Facts
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Bacterium of 2501 identified strains, as of 2001. Many
different diseases are caused by more than 1,400 serotypes of
this bacteria genus.
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“Salmonella” derived from Dr. Salmon, a U.S. veterinary
surgeon, who discovered and isolated the strain enterica or
choleraesuis from the intestine of a pig in 1885.
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They are ingested orally by contaminated food or water.
Refrigeration prevents growth but does not kill bacteria.
Heating at 57-60°C or 134-140°F has shown to be effective in
killing the bacteria.
Optimal growth: 37°C or 98.6°F
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Disease-associated facts
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“Salmonellosis”: Any of several bacterial infections
caused by species of Salmonella, ranging from mild
to serious infections.
Two main kinds in humans: enteric fever (typhoid
and paratyphoid) and gastroenteritis (non-typhoidal).
The main feature for S. diseases is the Type III
Secretion System, a needle-like multi-protein
complex that is associated with transferring toxic
proteins to host cells.
Principal habitats in different types of Salmonella
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Their principal habitat is the intestinal tracts and
bloodstream of humans, and in the intestinal tracts of
a wide variety of animals.
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The WHO groups Salmonella into 3 types:
- Typhoidal (enteric) Salmonella
(example: S. typhi)
٠causes typhoid and paratyphoid fever
٠restricted to growth in human hosts
٠principal habitat is in intestinal tracts
and the bloodstream
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- Nontyphoidal Salmonella (example: S. enteritidis, S.
typhimurium)
٠prevalent in gastrointestinal tracts of a broad range of
animals, including mammals, reptiles, birds and insects.
٠cause a whole range of diseases in animals and humans,
mainly gastroenteritis.
٠usually transferred animal-to-person, through certain food
products: fresh meat, poultry, eggs and milk
- fruits, vegetables, seafood
٠house and exotic pets, contamination through contact with
their feces
- Salmonella mostly restricted to certain animals, such
as cattle and pigs; infrequently in humans; if these
strains do cause disease in humans, it is often
invasive and life-threatening.
Salmonella
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Overview
History and Epidemiology
Molecular Biology
Clinical
Weaponization
History of Salmonella
Some historical figures are believed to have been killed by
• Salmonella:
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Alexander the Great died mysteriously in 323 B.C. In 2001,
a group of doctors at the University of Maryland suggested that
S. was the cause of death, based on a description of Alexander’s
symptoms written by the Greek author Arrian of Nicomedia.
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Prince Albert, the consort of Queen Victoria, died of a
Salmonella infection in 1861. During the Victorian era, an
estimated 50,000 cases per year occurred in England.
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History
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Scholars working on the history of Jamestown, Virginia, believe that a typhoid
outbreak was responsible for deaths of over 6000 settlers between 1607 and
1624.
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Typhoid Epidemic in the Spanish-American War (1898)
- In all, 20,738 recruits contracted the disease (82% of all sick soldiers), 1,590
died (yielding a mortality rate of 7.7%)
- It accounted for 87% of the total deaths from disease.
- A significant number of these deaths actually occurred at training areas in the
southeastern United States.
History
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Typhoid outbreak in British camps
during the South African War (18991902)
- more soldiers suffered from typhoid
fever than from battle wounds.
- British troops lost 13,000 men to
typhoid, as compared to 8,000 battle
deaths.
- outbreak was largely due to unsanitary
towns and farms throughout Africa, and
polluted soil was washed into the
network of streams and rivers during the
rainy season.
Epidemic potential during a war
prominent because of the disposal
problems of men’s discharges.
History
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Similar problems of sanitation occurred in urban areas. Many historic
documents report about typhoid outbreaks in England:
- Most outbreaks that were reported could be traced back to unsanitary water
supplies or polluted milk supplies.
- Dr. William Budd (1811-1880): documented his observations, published
them in the Lancet; It was known then that polluted water can spread the
disease. Budd urged for more disinfection and water treatment
- reports show that in the nineteenth century, population seemed powerless
against this disease even though they knew it was perfectly preventable.
- with the introduction of piped and filtered water supplies in most urban
areas, its prominence as a cause of death had diminished.
Salmonella vaccine
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First preventive measure against Salmonella was discovered in 1896, as an
antityphoid vaccine was developed by the British surgeon Almroth Wright.
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Vaccine consisted of heat-denatured, rudimentary killed whole-cell
bacteria; said to be highly effective.
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Early wars: -Immunization known, but new
-the minimum dosage had not been clearly refined;
British War Office authorized it on a voluntary basis only;
most soldiers refused to be immunized because of
violent reaction following injection; possible contraction
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Urban outbreaks: opposition to any type of vaccination; a way around the
problem of sanitation and cleanliness. It was seen as a disease of
“defective civilization …due to defective sanitation”.
Salmonella vaccine
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Between 1904-1914, the vaccine had become
respectable, in the scientific as well as military world.
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Vaccine was successfully used during World War I to
reduce the number of soldiers who died of enteric
fever (S. typhi).
First typhoid inoculation, 1909
United States Army Medical
School
Bottling typhoid vaccine, 1944
Division of Biologic Products, U.S.
Army of Medical Department
Professional Service Schools
History in the U.S.
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“Typhoid Mary” Mallon was the first famous
carrier of typhoid fever in the U.S.
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Some individuals have natural immunity to
Salmonella. Known as “chronic carriers”, they
contract only mild or asymptomatic disease,
but still carry the bacteria in their body for a
long time. These cases serve as natural
reservoir for the disease.
Approximately 3% of persons infected with S.
typhi and 0.1% of those infected with nontyphoidal salmonellae become chronic carriers
which may last for a few weeks to years.
One such case was Mary Mallon, who was
hired as a cook at several private homes in the
new York area in the early 1900’s.
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History: Mary Mallon
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Mary Mallon caused several typhoid outbreaks, moving from
household to household, always disappearing before an
epidemic could be traced back to the particular household
Mary was working in. All together, she had worked for 7
families, with 22 cases of typhoid and one death.
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She was finally overtaken by the authorities in 1907 and
committed to an isolation center on North Brother Island, NY.
There she stayed until she was released in 1910, on the
condition that she never accept employment involving food
handling.
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But: She was found to work as a cook and to cause typhoid
outbreaks again. She was admitted back to North Brother
Island, where she lived until her death in 1938.
Recent outbreaks
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More recently reported outbreaks in the U.S. involve
different kinds of Salmonella strains, predominantly S.
enteritidis and S. typhimurium.
In 1985, a salmonellosis (S. typhimurium) outbreak
involving 16,000 confirmed cases in 6 states by low fat milk
and whole milk from one Chicago dairy.
Largest outbreak of food-borne salmonellosis in the U.S.
Investigations discovered that raw and pasteurized milk had
been accidentally mixed.
Oregon: Intentional Contamination
of Restaurant Salad Bars
In September of 1984, 10
area restaurants in The
Dalles, Oregon, were
involved with outbreaks of
S. typhimurium
Outbreaks
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January 2000: infant aged 1 month visited a clinic
with fever and diarrhea. A stool specimen yielded
Salmonella serotype Tennessee. One week before
illness onset, the infant's family moved into a
household that contained a bearded dragon (i.e.,
Pogona vitticeps).
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During June 2002, a child aged 21 months was
admitted to a hospital with fever, abdominal cramps,
and bloody diarrhea. Blood and stool cultures yielded
Salmonella serotype Poona (from pet Iguana).
Foodborne diseases
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WHO: in 2000 that globally about 2.1 million people
died of foodborne illness
in industrialized countries, about 30% of people
suffer from foodborne diseases each year; around 76
million cases occur each year, of which 325,000
result in hospitalization and 5,000 in death.
(WHO, 2002)
Why do foodborne diseases emerge ?
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Globalization of food supply: for example, multistate
outbreaks of S. Poona infections associated with eating
Cantaloupe from Mexico (2000-2002)
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Unavoidable introduction of pathogens into new geographic
areas: for example, vibrio cholerae introduced into waters off
the coast of southern U.S. by cargo ship (1991).
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Travelers, refugees and immigrants exposed to unfamiliar
foodborne hazards.
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Changes in microorganisms: evolution of new pathogens,
development of antibiotic resistance, changes in the ability to
survive in adverse environmental conditions.
Why do foodborne diseases emerge ?
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Changes in human population: population of highly
susceptible people is expanding; more likely to
succumb to bacterial infections.
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Changes in lifestyle: Great amount of people eat
prepared meals. In many countries, the boom in food
service establishments is not matched by effective
food safety education and control.
Relative Frequency of the disease in the U.S.
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Estimate: 2 to 4 million cases of salmonellosis occur in the
U.S. annually (reported and unreported). Salmonella accounts
for the majority of food poisoning cases in the U.S
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Latest numbers:
In 2002, a total of 32,308 cases were reported from health
laboratories in 50 states.
The national rate of reported isolates was 11.5 per 100,000
population. Shows decrease of 7% compared to 1992, slight
increase of 2% from 2001.
Epidemiology
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The most commonly reported serotypes, in history
and present:
- S. typhi
- S. enteritides and S. typhimurium
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The “top 20” serotypes accounted for 80% of all
isolates reported in the U.S. in 2001.
Top 15 Salmonella Serotype list in the U.S., 2001
Country, Institution,
Biological origin
U.S.A., Centers of
Disease Control,
Control and
Prevention-FDDB Epi,
2001, Human
Total
Serot
ped
31,675
Rank
Serotype
% of Total
Serotyped
Count
1
Typhimurium
6,999
22.1
2
Enteridites
5,614
17.7
3
Newport
3,158
10
4
Heidelberg
1,884
5.9
5
Javiana
1,067
3.4
6
Montevideo
626
2
7
Oranienburg
595
1.9
8
Muenchen
583
1.8
9
Thompson
514
1.6
10
Saintpaul
469
1.5
11
Paratyphi B
tartrate positive
466
1.5
12
Infantis
440
1.4
13
Braenderup
388
1.2
14
Agona
370
1.2
15
Typhi
343
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Epidemiology
S. typhi (typhoidal Samonella)
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Causes enteric fever
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Have no known hosts other than humans.
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Transmission through close contact with infected or chronic
carriers. While direct person-to-person transmission through
the fecal-oral route is rare, most cases of disease result from
digestion of contaminated food or water.
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Since improvements in food handling, piped and filtered water
supplies as well as water/sewage treatment have been made,
enteric fever has become relatively rare in developed
countries.
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However, typhoid fever is still a big health-problem in
developing countries.
The WHO estimates that there are worldwide about 16
million of clinical cases annually, of which about 600,000
result in death. In comparison, about 400 cases occur each
year in the U.S., and 70% of these cases are acquired while
traveling internationally.
Salmonella typhi in developing countries
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Contaminated water is a
common cause in the spread
of typhoid fever. At the
time of rain, the
contaminated surface water
further contaminates water
supplies.
Severity, Morbidity and
complication rate is much
higher than in Europe and
North America due to lack
of antibiotics supply, water
filtration and treatment,
sterilization of water and
sanitation.
S. Typhi in the U.S.
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Almost 30% of reported cases in the U.S. are domestically
acquired.
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Although most cases are sporadic, large outbreaks do occur.
For example, outbreak linked to contaminated orange juice
in N. Y., caused by a previously unknown chronic carrier
(1991).
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Multi-drug resistance:
recent trend toward an increased incidence of multi-drug
resistant S. typhi in developing countries is reflected by
increase in the proportion of U.S. cases: 0.6% in 1985-1989 to
1.2% in 1990-1994.
Epidemiology
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S. enteriditis and typhimurium (non-typhoidal S.):
- are the 2 top serotypes in the U.S. since 1980’s
- cause gastroenteritis following ingestion of the
bacteria on or in food or on fingers and other objects
- cause the majority of cases of zoonotic salmonellosis
in many countries.
Humpty Dumpty
Salmonella Enteritidis
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transmitted to humans by
contaminated foods of
animal origin,
predominantly eggs. Raw
eaten or undercooked eggs
that have been infected in
the hen’s ovaries can cause
gastroenteritis
by R. Wayne Edwards
January 1999
Humpty Dumpty lay
on the ground
A crushed and
broken fella.
No one wanted
to put him
together
'Cause he had
salmonella.
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During the 1980s, illness related to contaminated eggs occurred most
frequently in the northeastern United States, but now it is increasing in
other parts of the country as well.
Salmonella Enteritidis Infections, United States, 1985–1999
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CDC, 2002: In the Northeast, approximately one in
10,000 eggs may be internally contaminated; one in
50 average consumers could be exposed to a
contaminated egg each year.
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In 1995: high of 3.9 per 100,000 population,
In 1999: 1.98 per 100,000, rate still decreasing due to
prevention and control efforts by the government.
S. typhimurium
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has been reported increasingly frequently as the cause
of human and animal salmonellosis since 1990, due to
antibiotic resistance
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Predominant multi-drug resistant strain DT 104,
which initially emerged in cattle in England, 1988
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In 1997, the WHO stated that some countries in
Europe had a staggering 20-fold increase in
incidences between 1980 and 1997, and a 5-fold
increase in the U.S. between 1974 and 1994, due to
antibiotic resistant strains
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intensive animal maintenance.
Epidemic measures
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Salmonellosis is a reportable disease.
An intensive search should be conducted for the source of an
infection and for the means (food or water) by which the
infection was transmitted.
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Samples of blood can be taken immediately for confirmation
and for testing for antibiotic sensitivity.
Samples of stool or urine may be taken after one week of onset
for effective confirmation.
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Food and water samples should be taken from suspected
sources of the outbreak. It is recommended to organize
temporary water purification and sanitation facilities until
longer term measures can be implemented.
Cost Estimates
The cost per reported case of human salmonellosis
range from US $100 to $1300 in North America and
Europe.
 The costs associated with individual outbreaks in
North America and Europe range from around
$60,000 to more than $20 Million.
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The total annual cost in the U.S. is estimated a total of
almost $400 Million.
Salmonella
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Overview
History and Epidemiology
Molecular Biology
Clinical
Weaponization
Salmonella Microbiology
Classification
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Enterobacteria
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Gram-negative
Facultative anaerobes
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Glucose-fermenting
Straight, rod
2-3 µm in length
Flagellated
Many serovars
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Typhi
Typhimurium
Enteriditis
LPS on Surface
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Lipopolysaccharide
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Protective outer layer of most strains
(not S. typhi)
Coded for by rfb locus on chromosome
Lipid core of LPS highly conserved across
serovars, but polysaccharide side chains are
highly polymorphic (nature of rfb gene)
LPS (cont.)
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Memory immune response and antibodies
directed against LPS
Polymorphic nature of side chains is
advantageous for bacteria
Since Typhi has outer capsule, this infection is
worse.
Infection
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Ingestion of contaminated
food or water
Passes through mucosa of
intestine to epithelial cells
Causes membrane ruffling
Releases effector proteins
through Type III Secretion
system
Endocytosis
Salmonella Entry
Membrane Ruffling
Virulence Factors
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Genes for virulence factors cluster in
pathogenicity islands (PI)
Genes acquired through lateral transfer
Bacteriophage and transposon insertion
sequences flank PI
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Maybe vehicles for transfer of PI to Salmonella at
one time
Acquisition of PI enhances virulence of
bacteria
Horizontal Transfer
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Transformation
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Conjugation
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Uptake of naked DNA
Mediates exchange of any part of DNA
F+ to FRequires cell to cell contact – conjugation bridge
Transduction
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Transfer of DNA by a phage
New phage: viral coat with bacterial DNA
Salmonella Pathogenicity Islands
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Salmonella
Pathogenicity Island 1
(SPI-1)
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entry into intestinal
epithelium
Enables pathogen to
exploit host intestinal
environment
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Salmonella
Pathogenicity Island 2
(SPI-2)
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intracellular bacterial
replication and initiation
of systemic infection
Do not influence
enteropathogenesis to
any great extent
Type III Secretion System (TTSS)
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Main way Salmonella
delivers virulence
factors to host
Made up of 20 proteins
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Assemble in step-wise
order
PrgI is a needle
structure extended by
protein base, forms a
channel to host
PrgI
Salmonella-host Interaction
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Two forms of TTSS
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SPI-1 TTSS probably causes initial interaction
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One encoded on SPI-1, other on SPI-2
Starts bacteria-mediated endocytosis
Entry activates SPI-2 TTSS to cause thorough
infection
Membrane Ruffling
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Cytoskeleton-associated proteins relocate to
site of bacterial entry
Bacterial effector proteins trigger cytoskeleton
rearrangements
Apical membrane surface undergoes structural
changes, resembling ruffling
This triggers endocytosis into vesicles
Slightly different from receptor-mediated
endocytosis
Salmonella Containing Vesicle
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After ingestion, Salmonella enters a SCV
through bacteria-mediated endocytosis
Lives and multiplies in SCV
Very little known about SCV or how bacteria
exist inside
A method to avoid host immune response
Phagosome: maturing SCV
SPI-1 Effector Proteins
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SipA
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Binds actin and stabilizes
filaments
Allows actin to
polymerize more easily
Maximizes efficiency of
Salmonella invasion
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SipC
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Aides in entry of other
SPI-1 effector proteins
Activtes G-actin to form
F-actin, then polymerize
Aides in cytoskeleton
rearrangements in
membrane ruffling
SopB
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Main virulence factor
Encoded by SPI-5
An enterotoxin associated with SPI-1 TTSS
Induces an increase in concentration of cellular
inositol polyphosphate
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Increased chloride secretion into lumen
Na+ follows to balance charge
Water follow to balance osmolarity
diarrhea
SPI-2 TTSS
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Activated once bacteria enters cell
Necessary for systemic infection
SPI-2 TTSS secretes effector proteins from
phagosome into cytosol
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Interfere with maturation of phagosome
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No fusion with lysosome
How Salmonella avoids degredation in cell
Flagella
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Another antigen
Host cytotoxic T-cell
response directed
against flagellar
epitopes
N- and C- termini are
highly conserved
Middle of flagellum is
variable
Phase I / II Flagella
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Operon encoding Phase I flagella also encodes
for a protein that represses trascription of
Phase II
The switch mediated by an enzyme that
inhibits Phase I, allowing Phase II
May help Salmonella avoid cell-mediated
immune response
Tumor Necrosis Factor-α
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Flagella from S. Typhimurium induces
expression of TNF-α through cell-mediated
reponse
Phase II flagella are less-potent inducers
Switching mechanism may provide bacteria
with a way to down-regulate inflammatory
response within host
Immune Response
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White blood cells recognize – trigger T cells, B cells
Two types of B cells: one to attack now, one for
memory
Macrophages and neutrophils attack bacteria, secrete
interleukins, causing cell-mediated response by Tcells
Antibodies from B cells attach to bacteria, allowing
cytotoxic T cells, macrophages, and neutrophils to
kill the organism
Inside Macrophages
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SPI-2 TTSS works in macrophages as well
Bacterium produces enzymes that inactivte toxic
macrophage compounds
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Homocysteine (Nitric Oxide antagonist)
Superoxide dismutase (inactivates reactive peroxides)
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Salmonella must produce additional factors to survive
limited nutrient base
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Allows bacteria to travel throughout body, causing
systemic infection (only in S. typhi)
Septicemia
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Invasion of bloodstream
spv genes causes
detachment of cells to
ECM and apoptosis
Spreads infection
Bacteria can enter
bloodstream and
lymphatic system
Main cause of death by
Salmonella
How do we respond?
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Microbiological view
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Vaccines
Dam
Antibiotics
Salmonella Vaccine Strategy
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Delete chromosomal regions that code for independent and
essential functions. This results in:
- low probability of acquiring both traits
- both traits:
* aro genes: aromatic compound biosynthesis
* pur genes: purine metabolism biosynthesis
Deletion strains
- can be grown on complete medium in lab
- in vivo, growth is reduced
- only a low level of infection is established
- immune system can mount a response
Vaccine suitable for humans and mice, chickens, sheep, cattle
DNA adenine methylase (Dam)
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Enzyme that methylates specific adenine
residues in Salmonella genome
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Regulates expression of about 20 bacterial
genes active during infection
Dam (-) mutants are not virulent
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Disrupts regulation of DNA replication and repair
Good antimicrobial potential
Current “hot topic” of research
Antibiotics
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Antibiotics are selective poisons
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Do not harm body cells
Target different aspects of bacteria, such as
ability to synthesize cell wall, or metabolism
MIC: Minimum Inhibitory Concentration
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the minimum amount of agent needed to inhibit the
growth of an organism
Antibiotic Resistance
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Bacteria can counteract antibiotics by:
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Preventing antibiotic from getting to target
Changing the target
Destroy the antibiotic
Bacteria can acquire resistance
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Horizontal transfer from another bacteria
Vertical transfer due to natural selection
Salmonella
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Overview
History and Epidemiology
Molecular Biology
Clinical
Weaponization
How Do You Catch Salmonella?
Food borne
 Transmitted via improperly prepared,
previously contaminated food or water
- Meat: poultry, wild birds, pork
- Dairy: eggs
 Pet turtles and lizards

How does Salmonella affect the
body?
Three clinical forms of salmonellosis
 - Gastroenteritis (S. typhimurium)
 - Septicemia (S. Choleraesius)
 - Enteric Fevers (i.e. S. typhi – Typhoid Fever)

Who Can Be Infected?
Everyone
 Especially: the elderly, infants,
immunocompromised patients (AIDS, sickle
cell anemia)

Factors Increasing Susceptibility
Identification I
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Laboratory identification of genus
Salmonella: biochemical + serological
tests
HOW?
- stool or blood specimens are plated on
agar media (bismuth sulfite, green agars,
MacConkey)
Suspect colonies further analyzed by
triple sugar iron agar/ or lysine-iron agar
- confirmed by antigenic analysis of O
(somatic) and H (flagellar) antigens Test
for antigens:
Identification II
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Use phenol red test:
- testing for lactic acid production
- if negative, diagnose (presence of red spots surrounded by a bright red
zone)
Salmonella typhimurium
Nontyphoidal Salmonella
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General Incubation: 6 hrs-10 days; Duration: 2-7 days
Infective Dose = usually millions to billions of cells
Transmission occurs via contaminated food and water
Reservoir:
a) multiple animal reservoirs
b) mainly from poultry and eggs (80% cases from eggs)
c) fresh produce and exotic pets are also a source of contamination (> 90% of
reptile stool contain salmonella bacterium); small turtles ban.
General Symptoms: diarrhea with fever, abdominal cramps, nausea and
sometimes vomiting
Nontyphoidal Salmonella
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Caused by S. typhimurium and S. enteritidis
Rainy season of tropical climates; Warm season of temperate climates
Growing rapidly in the U.S.: five-fold increase between 1974-1994
Centralization of food processing makes nontyphoidal salmonellosis
particularly prevalent in developing countries
Resistance is a concern, especially with multi-drug resistant S.
Typhimurium known as Definitive Type 104 (DT 104)
Nontyphoidal Salmonella:
Gastroenteritis
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Incubation: 8-48 hrs ; Duration: 3-7 days for diarrhea & 72
hrs. for fever
Inoculum: large
Limited to GI tract
Symptoms include: diarrhea, nausea, abdominal cramps and
fevers of 100.5-102.2ºF. Also accompanied by loose, bloody
stool; Pseudoappendicitis (rare)
Stool culture will remain positive for 4-5 weeks
< 1% will become carriers
Nontyphoidal Salmonella:
Bacteremia and Endovascular
Infections
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5% develop septicemia; 5-10% of septicemia patients develop localized
infections
Endocarditis: Salmonella often infect vascular sites; preexisting heart valve
disease risk factor
Arteritis: Elderly patients with a history of back/chest + prolonged fever or
abdominal pain proceeding gastroenteritis are particularly at risk.
- Both are rare, but can cause complications that may lead
to death
Septicemia
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Serotype S. choleraesius causes septicemia:
- prolonged state of fever, chills, anorexia, and anemia
- lesions in other tissues
- septic chock, death
Incidence of S. Enteritidis
Nontyphoidal Salmonellosis:
Localized Infections
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INTRAABDOMINAL INFECTIONS:
Rare, usually manifested as liver or spleen abscesses
Risk factors: hepatobiliary, abdominal abnormalities, sickle cell disease
Treatment: surgery to correct anatomic damages and drain abscesses
CENTRAL NERVOUS SYSTEM INFECTIONS:
Usually meningitis (in neonates, present with severe symptoms e.g. seizures,
hydrocephalous, mental retardation, paralysis) or cerebral abscesses
PULMONARY INFECTIONS:
Usually lobar pneumonia
Risk factors: preexisting lung abnormalities, sickle cell disease, glucocorticoid
usage
Typhoidal Salmonellosis: Enteric
Fever
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Incubation: 7-14 days after ingestion; Duration: several days
Infective Dose = 105 organisms
Symptoms:
a) 1st week: slowly increasing fever, headache, malaise, bronchitis
b) 2nd week: Apathy, Anorexia, confusion, stupor
c) 3rd week: rose spots (1-2 mm diameter on the skin): duration: 2-5 days,
variable GI symptoms, such as abdominal tenderness (majority), abdominal
pain (20-40% of cases) and diarrhea; enlargement of the spleen/liver, nose
bleeds, and bradycardia
neuropsychiatric symptoms: delirium and mental confusion
Long term effects: arthritis
Typhoidal Salmonellosis
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Late stage complications include intestinal perforation and
gastrointestinal hemorrhage
Immediate care such as increase antibacterial medications or
surgical resection of bowel
Other rare complications include inflammation of the pancreas,
endocardium, perocardium, myocardium, testes, liver, meninges,
kidneys, joints, bones, lungs and parotid gland and
hepatic/splenic abscesses
In general, symptoms of paratyphoid fever are similar to typhoid
fever, but milder with a lower mortality rate
Majority of bacteria gone from stool in 8 weeks; However, 1-5%
become asymptomatic chronic carriers: gallbladder is the
primary source of bacterium
Typhoidal Salmonella
Chest PA view shows pleural effusion, left lower pulmonary lobe atelectasis, medial and downward shift
of bowel gas, and an increase in the air-fluid level in the abdomen
Pictures
(A)
(A) In sub-acute infections, multiple white to yellow foci occur in the liver, spleen is enlarged, and
mesenteric lymph nodes may be enlarged
(B) Histopathological examination may reveal necrotizing splenitis and hepatitis, with necrotic
foci often accompanied by colonies of bacteria (arrow in right photo).
(B)
Treatment of Typhoidal
Salmonellosis
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Third generation cephalosporins or quinolones is the current treatment
IV or IM ceftriaxone (1-2g) is also prescribed; usually 10-14 days (5-7
days for uncomplicated cases)
Multi Drug Resistant (MDR) strains of S. typhi: quinolones are the
only effective oral treatment
Nalidixic acid resistant S. typhi (NARST) must be tested for sensitivity
to determine course of treatment
Sever typhoid fever (altered consciousness, septic shock):
dexamethasone treatment
Chronic carriers: 6 weeks of treatment with either oral amoxicillin,
ciprofloxacin, norfloxacin
Surgical intervention to remove damaged cells
Prevention
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Typhoidal S.:
- Generally treated with antibiotics
- vaccinations available; the CDC currently recommends
vaccination for persons traveling to developing countries
- Education of general public, especially in developing
countries; identification of all carriers and sources of
contamination of water supplies
- avoid risky foods & drinks:
buy bottled water or boil water for at least 1 minute;
COOK and CLEAN food thoroughly, avoid raw vegetables
and fruits
- WASH YOUR HANDS WITH SOAP AND WATER!!!
Preventive measures for non-typhoidal S.
- pasteurization of milk-products; Eggs from known
infected commercial flocks will be pasteurized
instead of being sold as grade A shell eggs.
- tracebacks, on-farm testing, quality assurance
programs, regulations regarding refrigeration,
educational messages for safe handling and cooking
of eggs
- Cross-contamination: uncooked contaminated foods
kept separate from cooked, ready-to-eat foods.
Salmonella Vaccines I
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Poultry vaccine: Megan™Vac 1
- applied to baby chicks via drinking water
and cattle. It stimulates immunity in the
chickens, preventing Salmonella infection
during the growing period which may
result in contamination and subsequent
food borne infection of humans
- targets S. Enteritidis
- Salmonella infection is stopped at lower
levels of the food chain will mean
increased productivity for the farmer and a
break in the cycle of Salmonella
transmission from animals à humans
Salmonella Vaccines II
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Today, three types of Typhoid Vaccines are available:
(1) inactivated whole-cell vaccine: 2 doses/ 4wks. Apart; single
booster dose recommended every 3 years
(2) Ty21a: a live, attenuated S. typhi vaccine. Administered orally (4
doses). Efficacy: 7 years
(3) Vi polysaccharide vaccine: from purified Vi polysaccharide from
S. typhi. Administered subcutaneously or intramuscularly. To
maintain protection, revaccination recommended every 3 years.
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These vaccines have been shown be 70-90% effective.
Salmonella
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Overview
History and Epidemiology
Molecular Biology
Clinical
Weaponization
CDC classification
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Category B agent: includes microorganisms that are moderately easy
to disseminate, have moderate morbidity (i.e., ability to cause
disease) and low mortality, but require enhanced disease
surveillance.
Biosafety Level 2
Risk Level 2: associated with human disease that is rarely serious
and prophylactic intervention is often available.
9 different species: Salmonella arizonae, cholerasuis, enteritidis,
gallinarum-pullorum, meleagridis, paratyphi (Type A,B,C), spp.,
typhi, and typhimurium
Salmonella typhi is the only species that requires import and/or
export permit from CDC and/or Department of Commerce; has high
droplet or aerosol production potential
WHO Global Salm Surv (GSS)
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GSS is an international Salmonella surveillance
program initiated in January 2002. It collects annual
summary data from member institutions all over the
world.
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The goal is to enhance the quality of Salmonella
surveillance, serotyping and antimicrobial resistance
testing and leading local interventions that reduce the
human health burden of Salmonella.
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A total of 138 laboratories were enrolled in the GSS
in September 2003.
Salmonella as a Bioterrorist Weapon: What states are
most at risk?
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The states most vulnerable to terrorist attack on the agricultural sector are
those with several or most of the following attributes:
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High density, large agricultural area
heavy reliance on monoculture of a restricted range of genotypes
major agricultural exporter, or heavily dependent on a few domestic
agricultural products
suffering serious domestic unrest, or the target of international terrorism, or
unfriendly neighbor of states likely to be developing BW programs
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First Use of Salmonella as a Bioterrorist Weapon
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From 1932-1945, Japan conducted biological warfare
experiments in Manchuria
At Unit 731, a biological warfare research facility, prisoners
were infected with Salmonella typhosa among other biological
agents
Additionally, a number of Chinese cities were attacked. The
Japanese contaminated water supplies and food items with
Salmonella. Cultures were also tossed into homes and sprayed
from aircraft
Due to inadequate preparation, training, and/or lack of proper
equipment, the Chekiang Campaign in 1942 led to about
10,000 biological casualties and 1,700 deaths among the
Japanese troops.
Oregon 1984: a religious cult known as the Rajneeshees, a Buddhist cult sought to run
the whole country by wining the local election in 1984 using salmonella bacteria. They
brewed a "salsa" of salmonella and sprinkled it on the town's restaurant salad bars. Ten
restaurants were hit and more than 700 people got sick.
• First large scale bioterrorism attack on American soil
• A communitywide outbreak of salmonellosis resulted; at least 751 cases were
documented in a county that typically reports fewer than five cases per year.
• Health officials soon pinned down salmonella as the cause of the sudden outbreak, but
put the blame on food handlers. In 1984, who could have imagined bioterrorism?
• caused by S. typhimurium as this type
Salmonella as a Bioterrorist Weapon
Wide distribution of food: contaminated food produced in one country can
cause illness in other countries
 Traceability
 Antimicrobial resistance: strains of
Salmonella are being found that have
multiple drug resistance
 Capacity building: Salm-gene project
used to enhance outbreak detection by
routinely sub-typing certain salmonellas
using molecular methods
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Salmonella as a Bioterrorist Weapon
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Contaminating unguarded food supplies
Some terrorist acts may be designed purely
to spread panic: contaminating the food
supply could bring economic and
agricultural production to a standstill
EX. If numerous food-borne outbreaks occurred
across the country, people would soon fear their
meals
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Unfortunately, people have reason to
worry: all these contaminations have
occurred naturally every year. If Mother
Nature can do this repeatedly, then a
terrorist should have no problem recreating
these outbreaks over and over in any
number of American cities.
Salmonella as a Bioterrorist Weapon
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readily accessible and easy to grow or make
Centralized food production: largely unmonitored food
supply; food that is tampered with can be widely + quickly
distributed
Terrorist groups could use infectious disease agents to
confuse public health officials into believing that outbreaks
are naturally occurring: it is estimated that 1.4 million
salmonella infections occur each year, but the CDC gets
reports of only about 38,000 annually
According to the Centers for Disease Control (CDC), only
32% of the reported outbreaks have a known etiology.
Salmonella as a Bioterrorist Weapon
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No food product is safe: vegetables and fruits are
the easiest to contaminate. Fresh-produce
wholesalers and distributors are notorious for
employing illegal immigrants and not checking
their background information.
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Even processed foods aren’t safe: Terrorists
could use heat-stable toxins that would survive
the packaging process.
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As more of our food becomes imported,
especially hard-to-clean off-season fruits and
vegetables, bioterrorists don’t even have to be
inside the United States to do damage
Salmonella as a Bioterrorist Weapon: Who might be
tempted to initiate an attack on the agricultural sector?
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Terrorist groups might be interested in agricultural bioweapons for a variety
of reasons:
1. international terrorist organizations: cause harm/injury to enemy states or
peoples
- in an ideologically-motivated terrorist attack there would be willing
assumption of responsibility by the perpetrator OR an attempt to disguise
the outbreak as natural.
2. Extreme activist groups:
- EX. anti-GMO groups for their potential value in deterring farmers from
the use of genetically engineered crops or animals
Salmonella as a Bioterrorist Weapon: What goals
might an attack on the agricultural sector serve?
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Food attack by a terrorist group: initiate point-source
epidemics using available Salmonella strains
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Destabilize a government by initiating food
shortages/unemployment: the potential for immense
economic damage due to contamination of the food supply
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Alter supply and demand patterns for a commodity:
an outbreak can trigger the imposition of trade restrictions. This
is turn would open up or close markets for others.
Salmonella as a Bioterrorist Weapon: What are the
special features of an attack on the agricultural sector?
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Salmonella is not hazardous to perpetrators: easy to produce, stockpile,
and disseminate
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Few technical obstacles to weaponization: it would not be difficult to
obtain Salmonella strains on the open market.
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Low security of vulnerable targets: Fields, supermarkets, restaurants
have essentially no security at all.
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Point source to mimic natural introduction: Because of the high
incidence of naturally-occurring diseases, a deliberately instigated
outbreak could be mistaken for a natural one
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Multiple point source outbreaks can be initiated by contaminating
imported feed or fertilizer, without even entering the country: allows
the possibility of initiating multiple outbreaks over a large geographic
area, in a way that mimics a natural event
Salmonella Dilemma
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Dissemination of genomic knowledge of salmonella can facilitate bioweapons development:
Alternative 1: Restrict dissemination of genomic knowledge
- short term: hinders development of a “super-Salmonella” terror
weapon
- long run: leaves us at the mercy of multi-drug resistant salmonella
strains ranging from incapacitating to lethal
Alternative 2: Disseminate genomic knowledge, but support
development of salmonella specific-drugs
- knowledge may provide a terrorist org. with the ability to develop
“super-Salmonella” terror weapons, but it provides us with the
opportunity to defend against all salmonella infection.
Combating Salmonella Bioterrorism
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Establish a national disease surveillance system that could not
only help uncover a terrorist attack but also recognize naturally
occurring outbreaks that now go undetected
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New technology: creating a diagnostic gene chip covering all
major diseases could give the health care provider instant
diagnoses. Similar gene chips could monitor the health of
livestock, poultry, and crops. Chips could be used during
various steps of food processing to ensure quality control and
food safety.
Lines of Defense
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Food processors should limit access to their production, storage and
packaging areas: rerouting traffic, installing locks
Randomized safety checkpoints: will increase fear of detection
COSTS:
Increase work force
Sampling and test costs
Record keeping
Government Action
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CDC monitors the frequency of Salmonella infections in the country
and assists the local and State Health Departments to investigate
outbreaks and devise control measures
FDA inspects imported foods, milk pasteurization plants, promotes
better food preparation techniques in restaurants and food
processing plants, and regulates the sale of turtles and it also
regulates the use of specific antibiotics as growth promotants in food
animals
USDA monitors the health of food animals, inspects egg
pasteurization plants, and is responsible for the quality of
slaughtered and processed meat.
EPA regulates and monitors the safety of our drinking water
supplies.
Biological Weapon Prevention

BTWC (Biological and Toxin Weapons Convention): drafted in 1972
- intended to prevent the development, production and stockpiling of
biological weapons
- pathogens or toxins in quantities that have no justification for protective
or peaceful services are to be eliminated
- today, 159 countries have signed the convention and 141 have ratified it
- however, more can be done: “ Factories in the former Eastern Europe
supply viruses that cause fatal diseases, such as E-Coli and Salmonella,
without checking the identities of the purchasers” (from the trials of the
largest fundamentalist org. in Egypt, Abu-al-Dahab)
Acknowledgements
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Dr. Geoffrey Zubay
Salwa Touma
Kathleen Kehoe