CAMPYLOBACTER & PLESIOMONAS
Ericka Mae P. Cruto Cru to BSMT 3-2
CAMPYLOBACTER
CAMPYLOBACTER
Campylobacter
The genus name Campylobacter was was derived from a Greek word for curved rod because they are curved, cur ved, spiral, S- shaped gramnegative bacteria (0.5 to 8 µm long and 0.2 to 0.5 µm wide). Most campylobacters campylobacters are asaccharolytic Although they they may appear to be strict anaerobes, they have have been grown in a microaerophilic environment. Most species are microaerophilic and majority appear to be pathogeniCampylobacter They have a single polar unsheathed un sheathed flagellum (monotrichous) or a flagellum at each end (amphitrichous). The motility of the bacteria is characteristically rapid and darting in corkscrew fashion.
Motility contributes to the ability ability of campylobacters campylobacters to colonize and infect intestinal mucosa. Multiplication of organisms organisms in the intestine leads to cell damag damage e and an inflammatory response. They They also have inability to ferment or oxidize oxidize the usual carbohydrate substrates available in the diagnostic laboratory. They reduce nitrates and they are also slowgrowing and fastidious.
Campylobacter spp. are microaerobic inhabitants of the gastrointestinal tracts of various animals, including poultry, dogs, cats, sheep, and cattle, as well as the reproductive organs of several animal species. Campylobacter spp. have been known to cause abortion in domestic animals, such as cattle, sheep, and swine and are primarily zoonotic organisms. Within the genus Campylobacter, Campylobacter, Campylobacter jejuni and Campylobacter coli are most often associated with infections in humans and are usually transmitted via contaminated food, milk, or water. Outbreaks have been associated with contaminated drinking water and improperly pasteurized milk.
Other campylobacters have been isolated from patients who drank untreated water, were compromised in some way, or were returning from international travel. The disease manifest most on infants and young adults, but all age is at risk. The transmission of campylobacterioses has been attributed to direct contact with animals and handling infected pets, such as dogs, cats, and birds, and indirectly by the consumption of contaminated water and dairy products and improperly cooked poultry. Person to person transmission has been reported, and some Campylobacter spp. are also sexually transmitted.
Campylobacters can cause either gastrointestinal or extraintestinal infections.
Extraintestinal disease including: • Meningitis • Endocarditis, • Septic arthritis
It is being recognized increasingly, particularly in patients with acquired immunodeficiency syndrome (AIDS) and other immunocompromised individuals. Gastroenteritis caused by Campylobacter spp. is usually a self-limiting illness and does not require antibiotic therapy.
Meningitis
Septic Arthritis
Campylobacter spp. produces three syndromes in humans: febrile systemic disease periodontal disease Gastroenteritis The most common clinical symptoms of Campylobacter infections (Campylobacteriosis) include: Diarrhea abdominal pain cramps fever headache nausea and vomiting
Gastroenteritis
Periodontal disease
Diagnostic Tool
There are no special requirements for the collection, transport, and processing of clinical specimens for the detection of campylobacters. Most common specimens submitted to the laboratory are feces and blood. Campylobacter spp. that cause enteric illness are isolated from stool samples and rectal swabs
Sometimes can be detected by direct Gram stain examination of stool. Subcultures from broths must be incubated in a microaerobic atmosphere or the organisms will not multiply. Turbidity is often not visible in blood culture media; therefore, blind subcultures or microscopic examination using acridine orange stain may be necessary.
The presence of Campylobacter spp. in blood cultures is detected effectively by carbon dioxide (CO2) monitoring.
Isolation from sources other than blood or feces is extremely rare but is ideally accomplished by inoculating the material (minced tissue, wound exudate) to a nonselective blood or chocolate agar plate and incubating the plate at 37° C in a CO2-enriched, microaerobic atmosphere.
Selective Media for the cultivation of Campylobacter Species
Campylobacter Selective (Skirrow) Agar
Campylobacter jejuni on a medium with blood, 48 hours, 42 °C, 10%CO2.
Biochemical test of Campylobacter spp. Species
Catal ase
Nitrate Reducti on
Urease
H2S Hippurat producti e on (TSI) Hydroly sis
Indoxyl Acetate Hydroly sis
C. Jejuni subsp. jejuni
+
+
-
-
+
+
C. coli
+
+
-
Variable result (V)
-
+
C. lari
+
+
-
-
-
-
C. fetus subsp.fetus
+
+
-
-
-
-
C. fetus subsp.vener ealis
+
+
-
-
-
-
GROWTH AT Species
15° C
25° C
42° C
C. Jejuni subsp. jejuni
-
-
+
C. coli
-
-
+
C. lari
-
-
+
C. fetus subsp.fetus
-
+
-
C. fetus subsp.venerealis
-
-
-
VIRULENCE FACTORS
FLAGELLA
IRON ACQUISITION
ADHESION & INVASION
CYTHOLETHAL DISTENDING TOXIN
Virulence Factor
Flagella – contributes to the bacteria’s motility
Adhesion and invasion – important factor for colonizing the host intestinal cells
Motility & chemotaxis – help lead the bacteria to its colonization site
Lipopolysaccharide (LPS) - plays a role in adherance as well as invading the immune system
Cytholethal distending toxin (cdt) – this toxin stops the cell’s growth cycle in G2 cdtA, cdtB and cdtC – genes that acitvates the cdt cdtB – can disrupt DNA in cell and causes cell cycle arrest
Iron acquisition – for sustaining nutrients w/in the host
Drug of Choice The drugs of choice for treating intestinal campylobacteriosis are:
Azithromycin
Erythromycin
Gentamicin Tetracycline, erythromycin, and chloramphenicol can be substituted for gentamicin.
Antimicrobial Resistance
Fluoroquinolones
Macrolides
Trimethoprim
Beta lactam antibiotics, including penicillin and most cephalosporins
Tetracycline
Quinolone
Kanamyci
Campylobacter species
Campylobacter fetus Campylobacter coli Campylobacter concisus Campylobacter curvus Campylobacter fetus subsp. fetus Campylobacter fetus subsp. veneralis Campylobacter gracilis Campylobacter helveticus Campylobacter hominis Campylobacter hyoilei Campylobacter hyointestinalis subsp. hyointestinalis Campylobacter hyointestinalis subsp. lawsonii Campylobacter insulaenigrae Campylobacter jejuni subsp. doylei
Campylobacter jejuni subsp. jejuni Campylobacter lanienae Campylobacter lari Campylobacter mucosalis Campylobacter rectus Campylobacter showae Campylobacter sputorum subsp. bubulus Campylobacter sputorum subsp. mucosalis Campylobacter sputorum subsp. sputorum Campylobacter upsaliensis
Several Campylobacter spp. have been implicated in human infection:
Campylobacter jejuni
Campylobacter coli
Campylobacter lari (laridis)
Campylobacter fetus contains two subspecies, Campylobacter fetus subsp. fetus and Campylobacter fetus subsp. venerealis.
Campylobacter jejuni
Known as the major infectious agent of human the most common cause of bacteria gastroenteritis most frequent cause of diarrhea C. jejuni can be isolated to patients with enteritis Majority of C. jejuni infections in humans is acquired during the preparation and eating of chicken highest infection rate of Campylobacter is during summer Infection with C. jejuni results in an acute inflammatory enteritis C. jejuni is thermophilic and capable of hydrolyzing hippurate. C. jejuni possess a gene coding for cytolethal distending toxin
Patient infected with C.jejuni present with a diarrheal disease that begins with mild abdominal pain within 2 to 10 days after ingestion of the organisms. Cramps and bloody diarrhea often follow the initial signs Patients may experience fever and chills and, rarely, nausea and vomiting. In most patients, the illness is self-limited and usually resolves in 2 to 6 days. Untreated patients can remain carriers for several month. Some cases patients are asymptomatic and mild, many complications have been reported in young children and immunocompromised patients, including bacteremia, hepatitis, cholecystitis, pancreatitis, abortion, myocarditis and meningitis. C. jejuni has been associated with Guillain-Barré syndrome
Diagnostic tool
Campy-BAP (blood agar plate), is a commonly used medium to isolate C. jejuni and other enteric campylobacters. This commercially available medium contains Brucella agar base, 10% sheep red blood cells, and a combination of antimicrobials — vancomycin, trimethoprim, polymyxin B, amphotericin B, and cephalothin. Positive hippurate hydrolysis is an important characteristic for the identification of C. jejuni. C. jejuni and other enteric campylobacters grow optimally at 42° C, growth of colon microbiota is inhibited at this higher temperature.
The typical colony morphology of C. jejuni and other enteric campylobacters is moist, runny looking, and spreading. Colonies are usually nonhemolytic; some are round and raised and others may be flat.
Blood-free, charcoal-based selective medium agar (CSM) Campylobacter jejuni
Campylobacter Blood Free Selective Medium (Modified CCDA)
Colony appearance of Campylobacter jejuni growing on Butzler agar, 48 hours, 42°C.
Campylobacter coli
Campylobacter coli was found more often in older patients and in patients having traveled abroad. C. coli infection is associated with acute enteritis and abdominal pain lasting for 7 days or more. The infections are generally self-limiting, complications may also involve bacteraemia, Guillain – Barré syndrome, reactive arthritis, and abortion. The primarily source of C. jejuni/ C. coli infections in human is caused while handling or during consumption of contaminated meat, especially poultry meat. Contact with pets and livestock, the consumption of contaminated water or raw milk and travelling in high prevalence areas are also considered risks factors in human disease.
They are non-spore formers and grow in micro aerobic conditions. C. coli grow slowly in culture and have an optimum temperature of 42°C, they do not grow at 25°C. Old cultures or ones exposed to air for extended periods tend to become spherical or coccoid.
C. jejuni and C.coli Primary Plating Media and Incubation time Modified Skirrow’s
media
Columbia blood agar base, 7% horse-lysed blood, and antibiotics (vancomycin, trimethoprim, and polymyxin B)
Campy-BAP
Brucella agar base with antibiotics (trimethoprim, polymyxin B, cephalothin, vancomycin, and amphotericin B) and 10% sheep blood
Blood-free, charcoal-based selective medium
Columbia base with charcoal, hemin, sodium pyruvate, and antibiotics (vancomycin, cefoperazone, and cyclohexamide)
Modified charcoal
cefoperazone deoxycholate agar (CCDA)
C. jejuni and C.coli Primary Plating Media and Incubation time Semi solid motility agar
Mueller Hinton broth II, agar, cefoperazone, and trimethoprim lactate
Campy-CVA
Brucella agar base with antibiotics (cefoperazone, vancomycin, and amphotericin B) and 5% sheep blood
Incubation time: 42° C under microaerophilic conditions* for 72 hr
Campylobacter fetus
Campylobacter fetus is a species of Gram-negative, motile bacteria with characteristic "S-shaped" rod morphology similar to members of the genus Vibrio. C. fetus is oxidase-positive. Campylobacter fetus can cause intestinal illness and, occasionally, severe systemic infections. Infections mainly affect persons at higher risk, including elderly and immunocompromised individuals, pregnant, or elderly persons and those with occupational exposure to infected animals. The reservoirs for C. fetus are mainly cattle and sheep. Products from these animals are suspected as sources for human infections. Campylobacter fetus is rarely isolated from food, causes severe infections, including bacteremia and meningitis.
C. fetus is an opportunistic human pathogen and can cause thrombophlebitis. It is the only Campylobacter species that can cause septicemia. Campylobacter fetus is devided into two subspecies: C. fetus subsp. venerealis C. fetus subsp. fetus
Campylobacter fetus subsp.fetus
Has been isolated from intestinal tracts of sheep and cattle and from tissues from sporadic abortions in these species Most cases of C. fetus infection are caused by C. fetus subsp. fetus. Campylobacter fetus subsp. fetus can be recovered from the intestinal tract of cattle and other animal species. Although C. fetus is primarily recognized as a veterinary pathogen, C. fetus subsp. fetus is occasionally diagnosed as an opportunistic emerging pathogen in humans. Infections usually occur in pregnant or immunocompromised individuals and are often systemic with a variety of neurological and vascular complications.
Campylobacter fetus subsp.fetus
Campylobacter fetus subsp.venerealis
Campylobacter fetus subsp. venerealis
C. fetus subsp. venerealis is restricted to cattle and causes bovine genital campylobacteriosis. C. fetus subsp. venerealis is associated with endemic abortion and fertility problems in certain areas. Although C. fetus subsp. venerealis has been isolated from humans, its role in human disease is uncertain. Bovine genital campylobacteriosis (BGC) is a venereal disease also known as bovine venereal campylobacteriosis (BVC). The causal agent of this sexually transmissible disease is Campylobacter fetus subsp. venerealis.
Campylobacter lari (C. laridis)
Campylobacter lari is a species of nalidixic acid-resistant, thermophilic, microaerophilic bacteria first isolated from human faeces. It shows anaerobic growth in the presence of trimethylamine N-oxide hydrochloride. It is commonly found in sea gulls.
In humans, it has been involved in cases of enteritis, severe abdominal pain and terminal bacteremia. Colonies are colorless, round and 1-1.5 mm in diameter after 72 hrs culture under microaerobic conditions on 5% blood agar. They do not hydrolyze hippurate.
C.lari can grow at 37 degrees celcius and 42 degrees celcius but not at 25 degrees Celcius. No growth in air at 25 or 37 degrees celcius.
(b) to (e) "C. laridis", showing different morphological forms of the cells
PLESIOMONAS
Plesiomonas
The genus Plesiomonas was formerly in the family of Vibrionacea but recent study shows that they are actually related to Enterobacteriacea. But unlike the Enterobacteriaceae it does not have the ability to produce gas from glucose, and it is the only oxidase-positive member. Plesiomonas is oxidase positive, glucose fermenting, facultatively anaerobic, gram negative bacilli Occur singly, in pairs, or in short chains or filamentous forms. They do not form spores or capsules and are motile by monotrichous or two to five lophotrichous flagella. Plesiomonas shigelloids is the only spp of the genus Plesiomonas
Plesiomonas Shigelloides
Found in soil and aquatic environments, isolates are generally found only in the fresh and estuarine waters of tropical and subtropical climates. They are widely distributed among warm- and cold-blooded animals, including dogs, cats, pigs, vultures, snakes, lizards, fish, newts, and shellfish. P.shigelloides can be a potential cause of enteric diseases in humans and it is isolated from a number of extraintestinal infections. The most common vehicle of transmission is the ingestion of contaminated water or food, particularly uncooked or undercooked seafood such as oysters, clams, or shrimp. They are generally 0.3-1.0 µm in width, 0.6-6.0 µm in length, motile with polar flagella, non-spore-producing, and facultatively anaerobic. Plesiomonas can be serotype by somatic O antigens and their H antigen.
Three major clinical types of gastroenteritis are caused by Plesiomonas:
The more common watery or secretory diarrhea A subacute or chronic disease that lasts from 14 days to 2 to 3 months A more invasive, dysenteric form that resembles colitis
The most common clinical symptom for all such patients is abdominal pain but patients also experience fever and vomiting. There are also reports of P. shegelloids infection in immunocompromised patiens. Occupational exposure can be a source of infections for veterinarians, zookeepers, aqua culturists, fish handlers, and athletes participating in water-related sports. Bactericimia and meningitis usually occurs in immunocompromised patiens and neonates.
Gastroenteritis is the disease with which P. shigelloides has been implicated. Clinical symptoms of gastroenteritis include: Fever Secretory or dysenteric diarrhea abdominal pain Vomiting, nausea, chills, arthralgia, and headache
Symptoms usually resolve spontaneously within two weeks but can last months. Asymptomatic cases are possible, but very rare.
Extraintestinal infections include:
Septicaemia
Meningitis
Osteomyelitis
Cellulites
septic arthritis
endophthalmitis
spontaneous bacterial peritonitis
acute cholecystitis
Diagnostic Tool
Plesiomonas spp. grow readily on most media routinely used in the clinical laboratory. After 18 to 24 hours incubation at 35° C, shiny, opaque, nonhemolytic colonies appear, with a slightly raised center and a smooth and entire edge. The easiest screening procedure is an oxidase test performed on colonies from nonselective media, such as SBA or CHOC agar.
Plesiomonas colonies are white to pink on this medium, and most coliform colonies are green or pink. P. shigelloides can be presumptively differentiated from similar genera with several key tests. The positive oxidase activity separates it from other Enterobacteriaceae, sensitivity to the agent O/129 separates it from Aeromonas, and its ability to ferment inositol separates it from all Aeromonas and almost all Vibrio spp. It can also be separated from the halophilic Vibrio spp. by its ability to grow in nutrient broth with 0%. NaCl and its inability to grow in nutrient broth with 6% NaCl.
Biochemical test of P.shigelloides Substrate or Property
Reaction
Oxidase
+
Indole
+
Urease
-
H2S
-
Simon Citrate
-
Lysine Decarboxylase
+
Beta-Galactosidase
+
Colonial Appearance and Characteristics
BA and Mac
Shiny, opaque,
smooth, non-hemolytic
Both NLF and LF
COL Agar
Antimicrobial Resistance
Although most cases of plesiomonad gastroenteritis are selflimiting, antimicrobial therapy is indicated for patients with severe or chronic disease. Similarly, extraintestinal infections, particularly among neonates, often require antimicrobial therapy. Studies have shown a general resistance to the penicillin class of antibiotics, but penicillins combined with a βlactamase inhibitor, as well as trimethoprimsulfamethoxazole, are active. There have been reports of resistance to more than one aminoglycoside (e.g., gentamicin, tobramycin, amikacin), but the quinolones, cephalosporins, and carbapenems appear to be effective therapy.
Drug of Choice
Ciprofloxalin
Virulence Factor
Unknown
