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Babylon university College of Medicine Microbiology Dept. Prof.Dr.Ilham AL-Saedi Enterobacteriaceae The Enterobacteriaceae is a large family of Gram-negative bacteria that includes many of the more familiar pathogens, such as Salmonella, Escherichia coli, Yersinia pestis, Klebsiella and Shigella. Other disease-causing bacteria in this family include Proteus, Enterobacter, Serratia, and Citrobacter. Characteristics of enterobacteriaceae: Members of the Enterobacteriaceae are rod-shaped, and are typically 1-5 μm in length. Like other proteobacteria, enterobacteria have Gram-negative stains, and they are facultative anaerobes, fermenting sugars to produce lactic acid and various other end products [lactose fermenters: produce pink-red colonies on MacConkey agar (Escherichia coli, Klebsiella, Citrobacter Enterobacter, Serratia. [nonlactose fermenters: pale-colour colonies on MacConkey agar (Salmonella, Shigella, Proteus). Most also reduce nitrate to nitrite. Most have many flagella used to move about, but a few genera are non-motile. They are not spore-forming. Catalase reactions vary among Enterobacteriaceae. Grow readily on MacConkey (MAC) and eosin methylene blue (EMB) agars 1 Grow readily at 35oC except Yersinia (25o-30oC) Motile by peritrichous flagella except Shigella and Klebsiella which are non-motile Do not form spores. Many members of this family are a normal part of the gut flora found in the intestines of humans and other animals, while others are found in water or soil, or are parasites on a variety of different animals and plants. Most members of Enterobacteriaceae have peritrichous, type I fimbriae involved in the adhesion of the bacterial cells to their hosts. Some enterobacteria produce endotoxins. Endotoxins reside in the cell cytoplasm and are released when the cell dies and the cell wall disintegrates. Some members of the Enterobacteriaceae family produce a systemic infection into the blood stream when all the dead bacterial cells release their endotoxins. This is known as endotoxic shock, and can be rapidly fatal. Identification of enterobacteriaceae: To identify different genera of Enterobacteriaceae, a microbiologist may run a series of tests in the lab. These include a range of tubes cultures and agar plates cultures such as: Phenol red Tryptone broth Phenylalanine agar for detection of production of deaminase, which converts phenylalanine to phenylpyruvic acid Methyl red or Voges-Proskauer tests depend on the digestion of glucose. The methyl red tests for acid 2 endproducts. The Voges Proskauer tests for the production of acetylmethylcarbinol. Catalase test on nutrient agar tests for the production of catalase enzyme, which splits hydrogen peroxide and releases oxygen gas. Oxidase test on nutrient agar tests for the production of the enzyme oxidase, which reacts with an aromatic amine to produce a purple color. Nutrient gelatin tests to detect activity of the enzyme gelatinase. In a clinical setting, three species make up 80 to 95% of all isolates identified. These are Escherichia coli, Klebsiella pneumoniae and Proteus mirabilis. Major Genera of enterobacteriaceae: Escherichia coli, Shigella, Salmonella, Citrobacter, Yersinia, Klebsiella, Enterobacter, Serratia, Proteus and Morganella. Natural Habitats of enterobacteriaceae: Environmental sites (soil, water, and plants) Intestines of humans and animals Modes of Infection of enterobacteriaceae: Contaminated food and water (Salmonella spp., Shigella spp., Yersinia enterocolitica, Escherichia coli O157:H7) Endogenous (urinary tract infection, primary bacterial peritonitis, abdominal abscess) Abnormal host colonization (nosocomial pneumonia) Transfer between debilitated patients Insect (flea) vector (unique for Yersinia pestis) 3 Enterobacteriaceae: Types of Infectious Disease: Intestinal (diarrheal) infection Extraintestinal infection - Urinary tract (primarily cystitis) - Respiratory (nosocomial pneumonia) - Wound (surgical wound infection) - Bloodstream (gram-negative bacteremia) - Central nervous system (neonatal meningitis) Urinary tract infection: Escherichia coli, Klebsiella pneumoniae, Enterobacter spp., and Proteus mirabilis Pneumonia: Enterobacter spp., Klebsiella pneumoniae, Escherichia coli, and Proteus mirabilis Wound Infection: Escherichia coli, Enterobacter spp., Klebsiella pneumoniae, and Proteus mirabilis Bacteremia: Escherichia coli, Enterobacter spp., Klebsiella pneumoniae, and Proteus mirabilis. Intestinal Infection: - Shigella sonnei (serogroup D) - Salmonella serotype Enteritidis - Salmonella serotype Typhimurium - Shigella flexneri (serogroup B) - Escherichia coli O157:H7 - Yersinia enterocolitica. Escherichia coli 4 E. coli is the head of the large bacterial family, Enterobacteriaceae, the enteric bacteria, is one of the most important enterobacteriaceae species; it is a gram negative rod, facultative anaerobic and non-sporulation. Usually motile. It can grow in media with glucose as the sole organic constituent. It produces polysaccharide capsule, positive test for indole, lysine decarboxylase and mannitol fermentation and produces gas from glucose. This bacterium is predominant among aerobic commensal bacteria in healthy human intestine. The bacterium can grow in the presence or absence of O2. Under anaerobic conditions it will grow by means of fermentation. E. coli uses mixed acid fermentation in anaerobic conditions, producing lactate, succinate, ethanol, acetate and carbon dioxide producing characteristic "mixed acids and gas" as end products. However, it can also grow by means of anaerobic respiration, since it is able to utilize NO3, NO2 or fumarate as final electron acceptors for respiratory electron transport processes. In part, this adapts E. coli to its intestinal (anaerobic) and its extraintestinal (aerobic or anaerobic) habitats. Antigenic structure of E.coli: somatic (O) antigen, capsular (K) antigen, flagellar (H) antigen. pilli: help in attachment and virulence: - bind to D-mannose residues on surface of cells. - Pyelonephritis associated pilli (pap). - Intestinal colonization factor antigen. 5 Virulence factors : - Adherence to uroepithelial cells by Pap pilli. - Capsule (K-antigen). - E.coli produce Exotoxins e.g. Hemolysins ,Enterotoxins causes diarrheas, Heat labile HL, Heat stable HS and Vero toxins like Shigella toxins. - Sidrophore-help survival of E.coli in iron-poor environment of human body fluids. Diseases caused by E. coli: 1. Urinary tract infection (UTI). -Urethritis: - Commonest cause (70%-90%). - More common in females due to shorter urethra. - 105 bacteria/ml of urine is significant - Common cause of hospital-acquired UTI due to urinary catheters. - Cystitis: (infection of bladder) Pain (dysuria) Frequency of maturation More common in females due to shorter urethra - Pyelonephritis: (infection of kidney) Fever (chills) Flank pain 2. Intestinal infections: a. Enterotoxigenic E. coli (ETEC): Virulence due to enterotoxins Act on small intestine Watery diarrhea (common couse of travelar diarrhea) Transmitted by contamination food and water -Produce Heat stable /Heat labile toxins 6 -Adheres to epithelium of small intestine. -Present with Nausea, Vomiting and Lose stool -H L like cholera toxin -Causes accumulation of fluids -Adhesive factors - Fimbriae specific receptor in the intestinal epithelium CFA -Mortality in children < 5 years b. Enteropathogenic E. coli (EPEC): Adhere to enterocytes, cause destruction of microvilli of small intestine. Infantile and childhood diarrhea (20% of bottlefed) Stool: watery, non-purulent, no blood. c. Enteroinvasive E. coli(EIEC): Cause invasion of enterocytes in large intestine Necrosis, ulceration and inflammation Stool: scanty, purulent and blood stain d. Enterohemorrhagic E. coli (EHEC) Due to verotxin –causes destruction of microvilli in large intestine. Produced by E. coli O157:H7 Hemorrhagic colitis with copious bloody stool without pus cells e. enteroaggragative E. coli EAggEC) f. Diffuse-aggregative E. coli. 3. Meningitis in newborns: form mothers genital tract (colonized with E.coli) 4. Opportunistic infections Peritonitis due to intestinal trauma 7 Wound infections Bacteremia gram-negative septic shock 5. Hospital-acquired infection: common cause Lab diagnosis of E. coli: - Specimens: urine, stool, pus - Culture on : MacConkey agar-lactose fermenter ENB agar- green metallic sheen Indole +, citrate – TSI: slant acid, butt –acid Treatment of E.coli diseases: - UTI: use antibiotics after C/S: Trimethoprim- Sulphamethoxazole). - Diarrhea: oral rehydration + ciprofloxacin - Meningitis: ceftriaxone (3rd generation cephalosporin). - Other diseases C/S: increasing resistance in E. coli 8