campylobacters, and helicobacter-

8/6/2019 Campylobacters, and Helicobacter-

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Dr.T.V.Rao MD

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Campylobacters causes Important

Zoonotic Infections

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First isolated as Vibrio fetusin 1909 from

spontaneous abortions in livestock Campylobacter enteritiswas not

recognized until the mid-1970s when

selective isolation media were developedfor culturing campylobacters from human

feces Most common form of acute infectious

diarrhea in developed countries; Higherincidence than Salmonella& Shigellacombined

History of Campylobacter

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Gram-negative Helical (spiral or curved) morphology; Tend to be

pleomorphic Characteristics that facilitate penetration and

colonization of mucosal environments (e.g.,motile by polar flagella; corkscrew shape)

Microaerophilic atmospheric requirements Become coccoid when exposed to oxygen or

upon prolonged culture Neitherferment nor oxidize carbohydrates

General Characteristics

Common to Superfamily

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Bacteria commonly found in animal feces. It

is one of the most common causes of humangastroenteritis in the world. Food poisoningcaused by Campylobacterspecies can beseverely debilitating, but is rarely life-

threatening. It has been linked withsubsequent development of Guillain-Barresyndrome (GBS), which usually developstwo to three weeks after the initial illness

Campylobacter jejuni

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Motility A Darting type

Distinctive rapiddarting motility

Long sheathedpolar flagellumat one (polar) orboth (bipolar)ends of the cell

Motility slowsquickly in wetmountpreparation

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Growth Requirements

Microaerophilic &capnophilic5%O2,10%CO2,85%N2

Thermophilic (42-43C)(except C. fetus)

Bodytemperature of

natural avianreservoir May become

nonculturable in nature

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Important Zoonotic

Infection Campylobacterremains the

leading cause worldwide ofzoonotic disease in humans,

surpassing Salmonellainfections. Moreover, casesare still believed to beunderreported. Oftencausing the same symptomsas Salmonella, such as mild tosevere diarrhea,

Campylobacterinfections canalso be an infectious triggerfor the more seriousGuillain-Barre Syndrome.

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C.jejuni carried in .Although C. jejuni is

not carried by healthyindividuals in the

United States orEurope, it is oftenisolated from healthycattle, chickens, birdsand even flies. It issometimes present innon-chlorinated watersources such as streamsand ponds.

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Feces refrigerated & examined within few hours

Rectal swabs in semisolid transport medium

Blood drawn for C. fetus

Care to avoid oxygen exposure

Selective isolation by filtration of stool specimen

Enrichment broth & selective media

Filtration:pass through 0.45 m filters

Laboratory Diagnosis

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Microscopy:

Gull-wing appearance in gram stain

Darting motility in fresh stool (rarely done inclinical lab)

Fecal leukocytes are commonly present

Identification:

Growth at 25o, 37o, or 42-43oC

Hippurate hydrolysis (C. jejuniis positive)

Susceptibility to nalidixic acid & cephalothin

Laboratory Identification

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An atmosphere with reduced O2 (5% O2)

with added CO2 (10% CO2)

At 42 (for selection)Several selective media can be used (eg,

Skirrows medium)

Two types of colonies:watery and spreading

round and convex

Culture

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Selective Medium

Blood-free,charcoal-basedselectivemedium agar(CSM) for

isolation ofCampylobacterjejuni

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Characteristic Result

Growth at 25 C -

Growth at 35-37 C -

Growth at 42 C +

Nitrate reduction +Catalase test +

Oxidase test +

Growth on MacConkey agar +

Motility (wet mount) +

Glucose utilization -

Hippurate hydrolysis +

Resistance to nalidixic acid -

Resistance to cephalothin +

Characteristics of C.jejuni

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http://en.wikipedia.org/wiki/Catalase_testhttp://en.wikipedia.org/wiki/Oxidase_testhttp://en.wikipedia.org/wiki/MacConkey_agarhttp://en.wikipedia.org/wiki/Hippuratehttp://en.wikipedia.org/wiki/Naladixic_acidhttp://en.wikipedia.org/wiki/Cephalothinhttp://en.wikipedia.org/wiki/Cephalothinhttp://en.wikipedia.org/wiki/Naladixic_acidhttp://en.wikipedia.org/wiki/Naladixic_acidhttp://en.wikipedia.org/wiki/Hippuratehttp://en.wikipedia.org/wiki/MacConkey_agarhttp://en.wikipedia.org/wiki/Oxidase_testhttp://en.wikipedia.org/wiki/Catalase_test


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Infectious dose and host immunity determine

whether gastro enteric disease develops Some people infected with as few as 500 organisms

while others need >106 CFU

Pathogenesis not fully characterized

No good animal model Damage (ulcerated, edematous and bloody) to themucosal surfaces of the jejunum, ileum, colon

Inflammatory process consistent with invasion of theorganisms into the intestinal tissue; M-cell (Payer's

patches) uptake and presentation of antigen tounderlying lymphatic system

Non-motile & adhesion-lacking strains are avirulent

Pathogenesis & Immunity

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Cellular components:

Endotoxin Flagellum: Motility

Adhesins: Mediate attachment to mucosa

Invasins

GBS is associated with C. jejuniSerogroup O19 S-layer protein microcapsule in C. fetus:

Extracellular components:

Enterotoxins Cytopathic toxins

Putative Virulence Factors

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Other Species of Campylobacters

C jejuni infections mayalso produce seriousbacteremic conditionsin individuals with

AIDS. Most reportedbacteremia's have beendue to Campylobacter

fetus fetus infection.

Campylobacter lari,which is found inhealthy seagulls, hasalso been reported toproduce mild recurrentdiarrhea in children

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Other Species of

CampylobactersCampylobacter

upsaliensismay causediarrhea or bacteremia,while Campylobacterhyointestinalis, whichhas biochemicalcharacteristics similar

to those of C fetus,causes occasionalbacteremia inimmunocompromised

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Gastroenteritis:Self-limiting; Replace fluids and electrolytes

Antibiotic treatment can shorten the excretion period;Erythromycin is drug of choice for severe or complicatedenteritis & bacteremia; Fluoroquinolones are highly active(e.g., ciprofloxacin was becoming drug of choice) but

fluoroquinolones resistance has developed rapidly sincethe mid-1980s apparently related to unrestricted use andthe use of enrofloxacin in poultry

Azithromycin was effective in recent human clinical trialsControl should be directed at domestic animal reservoirs

and interrupting transmission to humans

Treatment, Prevention & Control

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Complications are relatively rare, but infections have

been associated with reactive arthritis, hemolytic

uremic syndrome, and following septicemia,infections of nearly any organ. The estimatedcase/fatality ratio for all C. jejuni infections is 0.1,meaning one death per 1,000 cases. Fatalities are rare

in healthy individuals and usually occur in cancerpatients or in the otherwise debilitated. Only 20reported cases of septic abortion induced by C. jejunihave been recorded in the literature.

Complication are rare

but occurs occasionally

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Guillain-Barre syndrome (GBS), a demy elating disorder

resulting in acute neuromuscular paralysis, is a serioussequelae of Campylobacterinfection . An estimated one case ofGBS occurs for every 1,000 cases of Campylobacteriosis Up to40% of patients with the syndrome have evidence of recentCampylobacterinfection . Approximately 20% of patients withGBS are left with some disability, and approximately 5% diedespite advances in respiratory care. Campylobacteriosis is also

associated with Reiter syndrome, a reactive arthropathy. Inapproximately 1% of patients with campylobacteriosis, thesterile postinfection process occurs 7 to 10 days after onset ofdiarrhea . Multiple joints can be affected, particularly the kneejoint. Pain and incapacitation can last for months or become

chronic.

Sequelae to Infection

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Complications with

C.jejuni Guillain-Barre

Syndrome (GBS)

Favorableprognosis withoptimalsupportive care

Intensive-care unitfor 33% of cases

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A Tribute to Warren andMarshall

for Discovery of H.pylori

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Helicobacter pylori(H. pylori) is a type of bacteria.

Researchers believe thatH. pyloriis responsible for themajority of peptic ulcers.

H. pylori infection is common in the United States.About 20 per cent of people under 40 years old and halfof those over 60 years have it. Most infected

people, however, do not develop ulcers. WhyH.pylori does not cause ulcers in every infected person

is not known. Most likely, infection depends oncharacteristics of the infected person, the type ofH.pylori, and other factors yet to be discovered

Helicobacter pylori

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Helicobacter pylori is the prototype organism in this

group. It is associated with antral

gastritis,gastric ulcers, and gastriccarcinoma.

Helicobacter pylori

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Helicobacter pylori

Helicobacterpylori is a spiralgram negative

bacteria.It has a multiple

polar flagella

above the poleand motile

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Peptic ulcers have plagued men throughout the

centuries, but the exact cause of the condition was

uncertain. In 1940, Dr. A. Stone Freedberg ofHarvard Medical School identified unusual curvedbacteria in the stomachs of ulcer victims; hesuspected that they might be responsible for ulcersbut abandoned the research when his team wasunable to grow the bacteria in the laboratory

Beginning of Scientific

understanding

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Helicobacter pylori (H.pylori for short) was first

discovered in the stomachs of patients withgastritis & stomach ulcers nearly 25 years ago by Dr

Barry J. Marshall and Dr J. Robin Warren of Perth,Western Australia. At the time (1982/83) theconventional thinking was that no bacterium canlive in the human stomach as the stomach producedextensive amounts of acid which was similar in

strength to the acid found in a car-battery. Marshall& Warren literally re-wrote the text-books withreference to what causes gastritis & gastriculcers.

History of H.pylori

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Land Mark Changes in

H.pyloriThe name of thebacterium wasgrammaticallycorrected in 1987 toCampylobacter pyloriand,in1989 thebacterium was renamedHelicobacter pyloriandassigned as the typespecies of a novel genusdue to its 16s rRNAsequence.

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How is it transmitted?

Believed to be transmitted orally due to tainted foodor water.

Also believed to be passed through belching, orgastro-esophageal reflux, which is when smallamount of stomachs contents is forced up theesophagus.

After this process, its believed that the H. Pylori ispassed orally.


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Genes Contribute to

Pathogenicity

CAG Cytotoxinassociated gene

Vac Vacuolating

cytotoxin gene

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It is well established that urease, Vacuolatingcytotoxic VacA, and the pathogenicity island (cagPAI) gene products, are the main factors of virulence

of this organism. Thus, individuals infected withstrains that express these virulence factors probablydevelop a severe local inflammation that may inducethe development of peptic ulcer and gastric cancer.The way the infection spreads throughout the worldsuggests the possibility that there are multiplepathways of transmission.

Pathogenic Mechanism

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Culturing H.pylori

H.pylori grows onSkirrows mediumwith 1Vancomycin,2 Polymyxin3 Trimethoprim

Grows in 3 -6 days at370c

Colonies appearTranslucent 1-2 mm indiameter

Optimal growth occursin Microaerophicenvironment

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Biochemical Characters

Motile

Catalase +

Oxidase +

Strong producerof

Urease

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UreaseC=O(NH2)2 + H

+ + 2H2O HCO3- + 2 (NH4+)Urea Bicarbonate Ammonium

ionsAnd then

HCO3- CO2 + OH-

Urea Hydrolysis

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H.pylori is found in the deep mucus layer

Grows optimally at pH 6.0 to 7.0

But gastric mucosa has a strong buffering in spite oflower pH on the lumen side of stomach

H.pylori also produces a protease that modifies thegastric mucus and further reduces the ability of acid

through the mucus

Pathology and

Pathogenesis

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Mechanisms in

Pathogenicity

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Localization of H.pylori

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The potential character of H.pylori lie with

production of potent Urease activity which yields

production of Ammonia and further buffering acid.H.pylori is quite motile even in mucus finds its way

to epithelial surface

H.pylori overlies the gastric type but not intestinal

epithelial cells.

Pathogenesis

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Factors contributing to Peptic

ulceration There is a strong

association between

presence of H.pyloriinfection and pepticulceration

Mucosal inflammation

and damage involvesboth bacterial and hostfactors

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H.pylori causes Peptic ulcers in theStomach

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Lipopolysaccharides - damage mucosal cells

and Ammonia produced by Urease activity maydirectly damage cells.

Gastritis Chronic and active inflammationestablishes Polymorph nuclear and Mononuclearcell infiltration within the Epithelial and Laminapropria

Events lead to Destruction of epithelium iscommon.Glandular atrophy is common.

Factors influencing

Pathogenicity

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Acute infection

Upper Gastrointestinal illness

NauseaPain

Fever very occasionally

Acute symptoms lasts for < 1 week,May extend up to 2 weeks

Infection last for years, decades or even lifetime

Clinical Manifestations

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Consequences of H.pylori Infection

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About 90 % of patients with Duodenalulcer, and 50- 80 % of gastric ulcers areassociated with H.pylori infection.

H.pylori may have greater role inGastric carcinoma and Lymphomas

Association of Duodenal andGastric ulcers in H.pylori

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Specimens for histopathology Gastric

biopsy specimens can be used for

Histological examinationSpecimens obtained after Gastroscopy,

Biopsy, routine stains will demonstrate

Gastritis and special stains show curvedspiral organisms

Specimens collected in sterile saline mixedare used for culturing

Laboratory Diagnosis

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Endoscopy Gastric

Biopsy

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Serology The detection of

Antibodies in activeinfection is useful

But the tests are limited

utility as antibodiespersist even afterH.pylori infection iseradicated.

Several commercial kits

are available, but lacksthe role in identifyingacute infections.

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Special Tests for H.pylori

Rapid tests for detection ofUrease activity are widelyused in presumptiveidentification of GastricBiopsy specimens.

Gastric Biopsy can beplaced into urea containingmedium with colorindicator.

If H.pylori is present the

Urease rapidly splits ureaand resulting shift in pHyields a color change in themedium

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Urea Breath Test

H. pyloriinfection canbe detected in theexhaled breath using

this special test. Thistest is positive only ifthe person has acurrent infection.

Sensitivity andspecificity of this testranges from 94-98%.

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Carbon-14-urea Breath Test

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Urea Breath Test

In this test 13C or 14Clabeled urea isingested by patients

If H.pylori is presentthe urease activitygenerates labeledCo2 that can be

detected in thepatients exhaledbreath

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H.pylori and Cancer

. If a person has had an H.pyloriinfection constantly for 20-30years, it can lead to cancer of thestomach. This is the reason that

the World Health Organization's(WHO) International Agency forResearch into Cancer (IARC) hasclassified H.pylorias a Class- I-Carcinogen i.e. in the samecategory as cigarette smoking is

to cancer of the lung &respiratory tract.

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Triple therapy has prompt response, contain a

combination of following drugs

1 Metronidazole2 Bismuth subsalicylate or Bismuth sub citrate

3 Amoxicillin or Teracycles

administered up to 14 days

Eradicates H.pyloriIn 70 95 % of patients

Acid suppressing agent is supporting

Treatment

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Other alternatives

Proton pump inhibitor directly inhibit

H.pyloriCombined with

Amoxicillin

Clarithromycin or Amoxicillin

And Metronidazole

Other Drug Combinations

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A vaccination against Helicobacter pylori mayrepresent both prophylactic and therapeuticapproaches to the control of H. pylori infection.

Different protective H. pylori-derived antigens, suchas urease, vacuolating cytotoxin A, cytotoxin-associated antigen, neutrophil-activating protein andothers can be produced at low cost in prokaryote

expression systems and most of these antigens havealready been administered to humans and shown tobe safe.

Vaccines trails for H.pylori

are in Progress

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In Developed countries H.pylori are present in


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Barry J Marshall and J Robin


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Barry J. Marshall and J. RobinWarren have been awarded the

2005 Nobel Prize in medicine

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Helicobacter Foundation

The "HelicobacterFoundation" wasfounded by Prof. Barry

J. Marshall in early1994, and is dedicatedto providing you withthe latest information

about Helicobacter pylori,its diagnosis, treatmentand clinicalperspectives.

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Created by Dr.T.V.Rao MD for elearning resources for Microbiologists

in Developing World

Email

[email protected]

campylobacters, and helicobacter-

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