Microbiology SOLO 7 (Instructor's Version) PDF

MICROBIOLOGY SOLO 7 MacConkey Negative, Oxidase Variable Gram-Negative Bacilli and Coccobacilli, and Gram-Negative Bacilli that are Optimally Recovered on Special Media Presented By: Austin Reed, MS, MLS(ASCP)cm...

MICROBIOLOGY SOLO 7 MacConkey Negative, Oxidase Variable Gram-Negative Bacilli and Coccobacilli, and Gram-Negative Bacilli that are Optimally Recovered on Special Media Presented By: Austin Reed, MS, MLS(ASCP)cm 1 CHAPTER 31 Haemophilus species CHAPTER 31 ORGANISMS Haemophilus ducreyi Haemophilus haemolyticus Haemophilus influenzae  GENERAL CHARACTERISTICS Members of this genus are small, nonmotile, pleomorphic gram-negative bacilli Cells are typically coccobacillary or short rods The genus Haemophilus requires one of two factors, or even both, to grow in vitro X factor – Hemin V factor – Nicotine adenine dinucleotide (NAD) or NAD phosphate (NADP) Chocolate agar is the best agar to grow Haemophilus species because it contains both X and V factor BAP contains X factor but lacks V factor for optimal growth of Haemophilus species Haemophilus species can grow on BAP as well, under certain circumstances A satellite phenomenon can occur when Staphylococcus aureus grows alongside Haemophilus species and the Haemophilus will grow just around the Staphylococcus colonies – This happens because the Staphylococcus aureus lyses the red blood cells of the BAP on the streak line releasing X and V factor for Haemophilus to utilize EPIDEMIOLOGY OF HAEMOPHILUS DUCREYI Habitat Not part of normal human flora Only found in humans during infection Mode of transmission Person-to-person typically through sexual contact 5 VIRULENCE AND SPECTRUM OF DISEASE FOR HAEMOPHILUS DUCREYI Virulence factors Uncertain Capsule Pili Certain toxins are probably involved in cell attachment and penetration of host epithelial cells Spectrum of disease Chancroid Genital lesions progress from tender papules to painful ulcers with several satellite lesions Regional lymphadenitis is common 6  IDENTIFICATION OF HAEMOPHILUS DUCREYI Requires additional growth factors and special media for cultivation Mueller hinton with 1%IsoVitaleX and 3 µg/mL vancomycin BHI supplemented with 10% fetal bovine serus and 3 µg/mL vancomycin – Vancomycin is used to suppress the gram-positive growth of the genital tract, however, certain strains of H. ducreyi are susceptible to vancomycin so multiple media types are recommended Colonies are small, flat, and smooth and can be pushed intact across the agar surface (hockey puck) Requires X factor but does not need V factor Non-hemolytic Catalase negative Glucose fermentation variable Xylose, sucrose, and mannose fermentation negative PHYSEO USMLE: HAEMOPHILUS DUCREYI 3 2 1 8 EPIDEMIOLOGY OF HAEMOPHILUS HAEMOLYTICUS Habitat Normal human flora of the upper respiratory tract Mode of transmission Person’s own endogenous strain 9 VIRULENCE AND SPECTRUM OF DISEASE FOR HAEMOPHILUS HAEMOLYTICUS Virulence factors Uncertain Probably of low virulence Considered an opportunistic pathogen Spectrum of disease Associated with a wide variety of infections similar to H. influenzae 10  IDENTIFICATION OF HAEMOPHILUS HAEMOLYTICUS Colonies resemble H. influenzae on CAP except will be beta-hemolytic on rabbit or horse blood agar Requires both X and V factor for growth Beta-hemolytic Catalase positive Glucose fermentation positive Variable xylose fermentation Negative sucrose and mannose fermentation EPIDEMIOLOGY OF HAEMOPHILUS INFLUENZAE Habitat Normal human flora for the upper respiratory tract Mode of transmission Person to person contact Respiratory droplets Person’s own endogenous strain 12  VIRULENCE AND SPECTRUM OF DISEASE FOR HAEMOPHILUS INFLUENZAE Virulence factors Capsulated strains – Antiphagocytic – Additional cell envelope factors mediate attachment to host cells Unencapsulated strains – Pili and other cell surface factors mediate attachment Spectrum of disease Encapsulated strains – Meningitis – Epiglottitis – Cellulitis – Septic arhhritis – Pneumonia Unencapsulated strains – Localized infections – Otitis media – Sinusities – Conjunctivitis The most serious infections are caused by Haemophilus influenzae type b  IDENTIFICATION OF HAEMOPHILUS INFLUENZAE Unencapsulated strains are small, smooth, and translucent Encapsulated strains are larger and more mucoid Both types have a distinct mouse nest odor Requires X and V factors to grow Non-hemolytic Catalase positive Glucose and xylose fermentation positive Sucrose and mannose fermentation negative  PHYSEO USMLE: HAEMOPHILUS INFLUENZAE 5 2 1 6 7 4 3 15 ANTIMICROBIAL SUSCEPTIBILITY TESTING FOR HAEMOPHILUS SPECIES Haemophilus ducreyi Erythromycin is the drug of choice Other potentially active agents include ceftriaxone and ciprofloxacin Resistance to trimeth/sulfa and tetracycline has emerged Beta-lactamase-mediated resistance to ampicillin and amoxicillin is also known Haemophilus influenzae Usually ceftriaxone or cefotaxime for life-threatening infections For localized infections, several cephalosporins, beta-lactam/beta-lactamase inhibitor combinations, macrolides, trimeth/sulfa, and certain fluoroquinolones Beta-lactamase-mediated resistance to ampicillin is common Haemophilus haemolyticus Same as H. influenzae CHAPTER 32 Bartonella species CHAPTER 32 ORGANISMS Bartonella baciliformis Bartonella henselae Bartonella quintana  GENERAL CHARACTERISTICS The genus Bartonella grows primarily on CAP but will grow on BAP, just very slowly Typically the colonies will appear after 12 to 14 days and sometimes require up to 45 days The colonies will not grow on MAC There is no optimal procedure for the isolation of these organisms from clinical specimens This genus is short, gram-negative, pleomorphic rod-shaped, and very fastidious Oxidase and catalase negative EPIDEMIOLOGY OF BARTONELLA BACILIFORMIS Habitat Humans Mode of transmission Sandflies VIRULENCE FACTORS FOR BARTONELLA SPECIES Interact with host red blood cells, endothelial cells, and possibly bone progenitor cells Multiply and persist in the red blood cells Can cause angiogenic lesions Colonize the vascular endothelium Can inhibit endothelial cell apoptosis Activate monocyte and macrophage cells capable of producing potent angiogenic factors SPECTRUM OF DISEASE FOR BARTONELLA BACILIFORMIS Oroya fever Acute hemolytic bacteremia or a chronic vasoproliferative disease Carrion’s disease Produces a wart-like lesion called a verrugo Mortality rates range from 40-90% depending on host status and disease presentation EPIDEMIOLOGY OF BARTONELLA HENSELAE Habitat Domestic cats Mode of transmission Domestic cat bites or scratches Fleas and ticks  SPECTRUM OF DISEASE FOR BARTONELLA HENSELAE Associated with bacteremia, endocarditis, and bacillary angiomatosis Causes Cat Scratch Disease (CSD), rheumatic manifestations, and bacillary peliosis hepatitis, or splenic peliosis 22,000 cases of CSD and 80% of those cases are children Infection begins as a papule or pustule at the primary inoculation site, then develops to regional tender lymphadenopathy in 1-7 weeks. The infection can be chronic, self-limiting, or a severe systemic illness that can affect multiple body organs Diagnosis of CSD requires three of the four following criteria History of animal contact with evidence of a scratch or primary inoculation Negative laboratory studies for other causes of lymphadenopathy Characteristic histopathology of the lesion Positive skin test using antigen prepared from heat-treated pus collected from another patient’s lesion There are other Bartonella species that can cause CSD, but B. henselae is the most common cause of CSD EPIDEMIOLOGY OF BARTONELLA QUINTANA Habitat Humans Mode of transmission Human body louse and fleas  VIRULENCE AND SPECTRUM OF DISEASE FOR BARTONELLA QUINTANA Causes trench fever Mild influenza-like headache and bone pain to splenomegaly and a short-lived maculopapular rash Been reported in cases of bacteremia, endocarditis, chronic lymphadenopathy, and bacillary angiomatosis Primarily in patients infected with HIV  IDENTIFICATION OF BARTONELLA SPECIES Bartonella is extremely hard to cultivate in a routine microbiology laboratory Special media like BHI, fresh chocolate agar that has been prepared within at least 2 weeks, or heart infusion agars containing 5% fresh rabbit blood is needed to cultivate growth When Bartonella DOES appear on routine media, a small, irregular, dry, white “Cauliflower-like” colony that pits the agar will show up, usually on CAP Subbed Bartonella species will appear as tan and smooth The gram-stain appearance is small, slightly-curved, gram-negative rods Similar to Campylobacter, Helicobacter, or Haemophilus species These organisms are all Oxidase, urease, nitrate reductase, and catalase negative PHYSEO USMLE: BARTONELLA HENSELAE 7 6 5 3 1 2 4 28 ANTIMICROBIAL SUSCEPTIBILITY TESTING OF BARTONELLA SPECIES Treatment plans depend on the clinical manifestations of the disease and what species of Bartonella is isolated According to CLSI guidelines, Penicillin may not be an effective results of in vitro testing may not therapy despite susceptibility in vitro correlate with clinical efficacy Recent treatments with azithromycin indicate a successful, and more rapid resolution for the lymphadenopathy of CSD 29 CHAPTER 33 Campylobacter and Helicobacter species CHAPTER 33 ORGANISMS Campylobacter jejuni Helicobacter pylori  GENERAL CHARACTERISTICS These organisms require specialized media to isolate and identify Small, curved, gram-negative rods Most also require a microaerophilic atmosphere Campylobacter and Arcobacter Generally slow growing, fastidious, and asaccharolytic Helicobacter species Spiral-shaped organisms that resembled Campylobacter were isolated from the human stomach – Originally called Campylobacter pylori and has been changed to Helicobacter pylori Microaerophilic curved, helical, spiral, or fusiform gram-negative rods  EPIDEMIOLOGY OF CAMPYLOBACTER SPECIES Most Campylobacter species are pathogenic and associated with a variety of diseases in humans and other animals Inhabits GI tract of various animals Poultry Dogs Cats Sheep Cattle Usually transmitted via contaminated food, milk, or water Campylobacter jejuni infections are transmitted by direct contact during the preparation and eating of chicken in developed countries Highest rates of infection are late summer and early fall in the U.S.  SPECTRUM OF DISEASE: CAMPYLOBACTER SPECIES Causative agent of gastrointestinal or extraintestinal infections Extraintestinal diseases have been reported in patients with AIDS Meningitis Endocarditis Septic arthritis Gastroenteritis associated with Campylobacter species Is usually self-limiting and does not require antibiotic therapy Postinfection complications with C. jejuni have been recognized and include reactive arthritis and Guillain-Barré syndrome Acute demyelination of the peripheral nerves  IDENTIFICATION OF CAMPYLOBACTER SPECIES Successful isolation requires a selective media and optimal incubation conditions Campy BAP in a microaerophilic condition at 42°C up to 72 hours If Campylobacter is suspected in a blood culture, the bottle needs to be held for up to 2 weeks for detection Colonies are gray to pink or yellow gray and slightly mucoid Oxidase and Catalase Positive Unable to grow in 3.5% NaCl Growth at 42°C Hippurate Hydrolysis Positive for C. jejuni All other pathogenic Campylobacter are negative Nitrate reduction positive PHYSEO USMLE: CAMPYLOBACTER JEJUNI 5 3 1 4 5 8 7 2 6 9 36  EPIDEMIOLOGY OF HELICOBACTER PYLORI Primary habitat is the human gastric mucosa Distributed worldwide and acquired early in life in underdeveloped countries Mode of transmission is unknown Oral-oral, fecal-oral, and a common environmental source have been proposed, but there are no conclusions Capable of colonizing the mucous layer of the antrum and fundus of the stomach but does not invade the epithelium Motility allows it to escape the acidity of the stomach and burrow through and colonize the gastric mucosa  SPECTRUM OF DISEASE: HELICOBACTER PYLORI Helicobacter pylori produces urease that hydrolyzes urea-forming ammonia This creates a significantly higher pH around the site of infection and protects the organism from the gastric secretions Helicobacter pylori produces a protein called CagA and injects the protein into the gastric epithelial cells Affects host cell gene expression, altering cell structure and interactions with neighboring cells, enabling H. pylori to invade the gastric epithelium Positive antibody response to CagA protein are at an increased risk for developing both peptic ulcer disease and gastric carcinoma Colonization persists despite the host’s immune response  IDENTIFICATION OF HELICOBACTER PYLORI Gastric biopsy specimens that are preserved in 10% formaldehyde or paraffin- imbedded tissue are used for the diagnosis of an Helicobacter pylori infection A noninvasive test is the urea breath test Relies on the presence of Helicobacter pylori urease If the organism is present, a radioactively labeled urea will be hydrolyzed by the urease produced by the organism to form ammonia and labeled bicarbonate that will be exhaled Positive for oxidase, positive, and urease  PHSYEO USMLE: HELICOBACTER PYLORI 3 2 4 7 6 8 5 1 40 ANTIMICROBIAL SUSCEPTIBILITY TESTING FOR CAMPYLOBACTER JEJUNI AND HELICOBACTER PYLORI Campylobacter jejuni Susceptible to many antimicrobial agents including macrolides, tetracyclines, aminoglycosdies, and quinolones Erythromycin is the drug of choice for patients with severe gastroenteritis with signs of severe dehydration and/or bacteremia Fluoroquinolones was the therapy most commonly prescribed, however a rapidly increasing proportion of Campylobacter species have been identified as resistant to fluoroquinolones Helicobacter pylori Except metranidazole and clarithromycin, most susceptibility assays are unsuccessful in predicting clinical outcomes Readily becomes resistant when metronidazole, clarithromycin, azithromycin, rifampin, or ciprofloxacin is prescribed as a single agent Current regimens recommend a triple-drug therapy including – Proton pump inhibitor (prilosec or nexium) – Clarithromycin – Amoxicillin or metronidazole CHAPTER 34 Legionella pneumophila  GENERAL CHARACTERISTICS All Legionella species are obligate aerobes that grow best between 20-45°C They are faintly staining, thin gram-negative bacilli Does not grow on routine media and requires a medium supplemented with iron and L-cysteine for growth The organism utilizes protein instead of carbohydrates for their energy source 58 species belong to the Legionella genus, however, Legionella pneumophila dominates the genus as the predominant human pathogen  EPIDEMIOLOGY OF LEGIONELLA PNEUMOPHILA Legionellae are widely distributed in the environment Most individuals are exposed to Legionella species HOWEVER, few develop symptoms Primarily found in aquatic habitats and thrive at warmer temperatures Legionella pneumophila can survive up to 14 months in water with only a slight loss in viability Legionella is also widely distributed in man-made facilities Air conditioning ducts, cooling towers, potable water, large, warm-water plumbing systems, and humidifiers Infections are acquired exclusively from environmental sources No person-to-person transmission has been documented  VIRULENCE AND SPECTRUM OF DISEASE FOR LEGIONELLA PNEUMOPHILA Legionella can multiply within amoebae and certain host cells Can also multiply within biofilms This contributes to the organism’s survival in the environment Outer membrane proteins, macrophage infectivity potentiator, genes encoding for the type II secretion required for intracellular growth, Type IV pili, flagella, and a dot/icm type IV secretion system aid in the invasiveness of Legionella pnuemophila Legionnaires’ disease is the main disease state of Legionella pneumophila Severe pneumonia with a case fatality rate of 10-20% – Global health issue Pontiac fever Mild, self-limited, nonfatal, influenza-like respiratory infection Extrapulmonary infections can occur such as within wounds, abscesses, encephalitis, and endocarditis  IDENTIFICATION OF LEGIONELLA PNEUMOPHILA Because Legionella pneumophila is biochemically inert, definitive identification requires the facilities of a specialized reference laboratory There are ways we can get a presumptive identification Specimens suspect of Legionella pneumophila infection are planted to two plates: BCYE with and without L-cysteine Legionella pneumophila will only grow on the BCYE WITH L-cysteine Direct immunofluorescent antibody (DFA) can be used on respiratory secretions to test for the presence of Legionella pnuemophila If Legionella pneumophila is present, the fluorescent antibodies attach to the organism’s antigens and cause the organism to glow a bright green DIRECT IMMUNOFLUORESCENT ANTIBODY FOR LEGIONELLA PNEUMOPHILA 47 PHYSEO USMLE: LEGIONELLA PNEUMOPHILA 12 1 11 7 4 6 2 9 8 5 10 3 48 ANTIMICROBIAL SUSCEPTIBILITY TESTING FOR LEGIONELLA PNEUMOPHILA In vitro susceptibility studies are not predictive of clinical response Fluoroquinolones and a newer macrolide (clarithromycin and azithromycin) is typically used for Legionella pneumophila infections Penicillins, cephalosporins, and aminoglycosides should not be used 49 CHAPTER 35 Brucella species CHAPTER 35 ORGANISMS Brucella abortus Brucella melitensis Brucella suis Brucella canis  GENERAL CHARACTERISTICS Brucellae are small, facultative, intracellular, nonmotile, aerobic, gram-negative coccobacilli Requires CO2 for growth These organisms are all considered potential agents of bioterrorism Although most isolates will grow on BAP and CAP, more enriched agars and special incubation conditions are generally needed for optimal recovery Blood culture systems are typically the best way to facilitate growth for Brucella species  EPIDEMIOLOGY OF BRUCELLA SPECIES Brucella species occurs worldwide, espeically in the Mediterranean and Persian Gulf contries Capable of survival for extended periods Can survive in fresh cheese for several months Each of the 4 Brucella species have a preferred animal host Brucella abortus – cattle and buffalos Brucella melitensis – sheep, goats, or camels Brucella suis – swine and a variety or wild animals Brucella canis – dogs Humans become infected by four primary routes Ingestion of unpasteurized animal milk (Most common means of transmission) Inhalation of infected aerosolized particles (Most important source of transmission for laboratory acquired infections Direct contact with infected animal parts through ruptures of skin and mucous membranes Accidental inoculation of mucous membranes by aerosolization  VIRULENCE FACTORS AND SPECTRUM OF DISEASE FOR BRUCELLA SPECIES Able to exist both intracellular and extracellular environments Ingested by neutrophils, within which they will replicate and cause cell lysis Circulate in the bloodstream and phagocytized by mononuclear phagocytic cells in the spleen, liver, and bone marrow Brucella melitensis and abortus are the most virulent for humans Cause a condition called brucellosis which is considered a zoonotic infection Can range from asymptomatic to serious debilitating disease Most likely a systemic infection that can involve any organ in the body Symptoms – Fever and chills – Weight loss – Night sweats – Headache – Muscle aches – Fatigue – Depression – Lymphadenopathy and splenomegaly are common physical findings  IDENTIFICATION OF BRUCELLA ABORTUS Small, convex, smooth, translucent, gamma hemolytic, and slightly yellow after at least 48 hours of incubation May brown with age All Brucella strains will be catalase positive, urease positive and oxidase positive Differentiated by the rapidity of which the organism hydrolyzes urea and ability to produce H2S, and inhibition to aniline dyes Brucella abortus does not grow in ambient air and can be used to differentiate from the other Brucella species 2 hours for positive urease H2S produced Inhibited by thionine Not inhibited by fuchsin IDENTIFICATION OF BRUCELLA MELITENSIS CO2 not required for growth 2 hours for positive urease H2S negative Not inhibited by any dyes IDENTIFICATION OF BRUCELLA SUIS CO2 is not required for growth 15 minutes for a positive urease H2S is variable Is not inhibited by thionine Inhibited by fuchsin  IDENTIFICATION OF BRUCELLA CANIS CO2 is not required for growth 15 minutes for positive urease H2S is not produced Is not inhibited by thionine Inhibited by fuchsin Colonies are rough instead of smooth  PHYSEO USMLE: BRUCELLA SPECIES 2 1 4 3 59 ANTIMICROBIAL SUSCEPTIBILITY TESTING FOR BRUCELLA SPECIES Because of their fastidious nature, susceptibility testing is not reliable Brucellosis patients undergo a prolonged treatment for around 6 weeks with antimicrobials that can penetrate macrophages and act in acidic environments Doxycycline or tetracycline in combination with streptomycin or rifampin is recommended CHAPTER 36 Bordetella CHAPTER 36 ORGANISMS Bordetella pertussis Bordetella parapertussis GENERAL CHARATERISTICS Bordetella species are nonmotile and infect only humans These Bordetella species do not grow on MAC, unlike B. bronchiseptica 63  EPIDEMIOLOGY OF BORDETELLA PERTUSSIS AND PARAPERTUSSIS Before the introduction of the vaccine, pertussis (whooping cough) became an epidemic that cycled approximately every 2- 5 years Transmission occurs person-to-person through inhalation of respiratory droplets Humans are the only known reservoir VIRULENCE FACTORS AND SPECTRUM OF DISEASE FOR BORDETELLA PERTUSSIS AND PARAPERTUSSIS Adhesion (auto transporters) Pertussis toxin Enters the bloodstream, binds to specific receptors on host cells and disrupts several host cell functions Center membrane of B. pertussis blocks access of the host’s lysozyme to the bacterial cell wall via its outer membrane Clinical pertussis is usually a disease of children and can be divided into three symptomatic stages Catarrhal stage – symptoms are the same as for a mild cold with a runny nose and a mild cough Paroxysmal stage – Severe and violent coughing increase; associated with vomiting and “whooping” as the result of air rapidly inspired into the lungs past the swollen glottis Convalescent stage – Recovering stage  IDENTIFICATION OF BORDETELLA PERTUSSIS Selective media for suspected patients would be Bordet-Gengou and Regan-Lowe Bordetella species are small and faintly staining coccobacilli that appear singly or in pairs Catalse positive Oxidase positive Non motile Nitrate negative Urease negative Growth on Regan Lowe within 3-6 days No growth on BAP or MAC  PHYSEO USMLE: BORDETELLA PERTUSSIS 3 8 2 1 10 4 7 9 6 5 67  EPIDEMIOLOGY OF BORDETELLA PARAPERTUSSIS Catalase positive Oxidase negative Nonmotile Nitrate negative Urease positive (24 hours) Growth on Regan Lowe within 2-3 days Growth on BAP and variable growth on MAC ANTIMICROBIAL SUSCEPTIBILITY TESTING FOR BORDETELLA SPECIES Routine laboratories do not perform routine susceptibility testing Organisms remain susceptible to erythromycin or the new macrolides, ketolides, quinolones, and other antibiotics Three erythromycin-resistant strains have been discovered Resistant to most oral cephalosporins CHAPTER 37 Francisella tularensis  GENERAL CHARACTERISTICS Francisella tularensis is considered facultative, intracellular pathogens that require cysteine, cystine, or another sulfhydryl and a source of iron for enhanced growth Faintly staining, tiny, gram-negative coccobacilli Non-spore forming, strict aerobes Considered a bioterrorism agent  EPIDEMIOLOGY OF FRANCISELLA TULARENSIS Francisellaceae are widely distributed throughout the environment Francisella tularensis is the agent of human and animal tularemia Carried by many species of wild rodents, rabbits, beavers, and muskrats in North America Most cases in the United States are sporadic, occurring during the summer months, and seen in the states of South Dakota, Arkansas, Missouri, and Oklahoma Humans become infected be handling the carcasses or skin of infected animals, inhaling infective aerosols, or ingesting contaminated water Can also be infected by insect vectors such as the deerfly and tick  VIRULENCE FACTORS FOR FRANCISELLA TULARENSIS Capsule appears to be a necessary component for the full virulence Allows the organism to avoid immediate destruction by PMNs Extremely invasive and can survive in the cells of the reticuloendothelial system Humans can be infected by fewer than 50 organisms by either aerosol or cutaneous routes Typically takes other organisms hundreds or even thousands to infect  SPECTRUM OF DISEASE FOR FRANCISELLA TULARENSIS Tularemia is recognized worldwide and has also been called rabbit fever, deer fly fever, and market men’s disease Typical clinical presentation after inoculation is a lesion at the site of infection (bite or abrasion) and progresses to an ulcer. Lymph nodes adjacent to the site of inoculation are enlarged and necrotic. Once the organism enters the bloodstream, patients become systemically ill Tularemia can range from mild and self-limiting to fatal Tularemia exists as 6 different types Ulceroglandular – common with ulcers and lymphadenopathy Glandular – common with lymphadenopathy Oculoglandular – conjunctivitis and lymphadenopathy Oropharyngeal – Ulceration in the oropharynx Systemic (typhoidal) tularemia – Acute with septicemia. No ulcers or lymphadenopathy Pneumonic tularemia – Acquired by inhalation of infectious aerosols or by dissimination from the blood stream (the most serious form of tularemia  IDENTIFICATION OF FRANCISELLA TULARENSIS Colonies are transparent and mucoid Isolates should be sent to a reference laboratory for confirmatory testing and characterization Indications of a possible Francisella tularensis isolate Unusual gram stain with small, poorly staining gram- negative rods seen as single cells or amorphus gram- negative mass without distinct cell forms Subcultures yield primarily pinpoint colonies on CAP – Grows better on CAP than BAP Oxidase negative Weak catalase positive Small gram-negative rods are observed in a positive blood culture in which time of detection is longer than 24 hours PHYSEO USMLE: FRANCISELLA TULARENSIS 1 4 2 5 3 6 76 ANTIMICROBIAL SUSCEPTIBILITY TESTING FOR FRANCISELLA TULARENSIS No standardized susceptibility test exists Streptomycin is the drug of choice Gentamycin is a possible alternative Fluoroquinolones are promising for treatment of severe tularemia Development of antibiotic resistance is rare CHAPTER 38 Streptobacillus moniliformis  GENERAL CHARACTERISTICS Streptobacillus moniliformis is a gram-negative bacillus that requires media containing blood, serum, or ascites fluid as well as incubation under CO2 for isolation Considered a faculatative anaerobe that is highly pleomorphic  EPIDEMIOLOGY OF STREPTOBACILLUS MONILIFORMIS Natural habitat is the upper respiratory tract of wild and laboratory rats Mice, gerbils, squirrels, ferrets, and weasels Has been isolated from other animals such as cats and dogs that have fed on rodents Pathogenic for humans and is transmitted by two routes Rat bite – Or direct contact with rat feces or saliva Ingestion of contaminated food – Unpasteurized milk Occurs worldwide  VIRULENCE FACTORS AND SPECTRUM OF DISEASE FOR STREPTOBACILLUS MONILIFORMIS Virulence factors are unknown but is known to spontaneously develop L forms Bacteria without cell walls – Allows the bacteria to persist The disease state is called rat-biter fever or Haverhill fever Patients develop acute onset of chills, fever, headache, vomiting, and often severe joint pains Symptoms usually occur within 3-10 days after exposure Patients may develop a rash on the palms, soles of the feet, and other extremities Other complications can occur including endocarditis, septic arthritis, pneumonia, pericarditis, brain abscesses, prostatitis, and pancreatitis  IDENTIFICATION OF STREPTOBACILLUS MONILIFORMIS Streptobacillus moniliformis is a pleomorphic gram-negative rod Cells may appear strait or variable size, or as long, tangled chains and filaments with bulbar swellings May resemble a string of pearls Growth on BAP can occur after 48 hours of incubation at 37°C Colonies are embedded in the agar and have a “fried egg” appearance with a dark center and a flattened, lacy edge These colonies typically have transformed into the L form and a gram-stain will show bi-polar staining coccoid forms – Acridine orange may be used to confirm bacteria is present since a gram- stain relies heavily on the bacterial cell wall Indole negative Catalase negative Oxidase negative Nitrate reduction negative Nonmotile Urea Negative Lysine decarboxylase negative H2S is not produced in TSIA BUT can be detected using lead acetate paper ANTIMICROBIAL SUSCEPTIBILITY TESTING FOR STREPTOBACILLUS MONILIFORMIS No standard methods have been established Penicillin is considered the drug of choice Aminoglycosides or tetracycline can be used to eliminate the L forms or for patients allergic to penicillin BIOTERRORISM AGENTS CATEGORY A BIOTERRORISM AGENTS Category A – highest priority agents Certain bacteria are considered bioterrorism agents because of Easily disseminated or transmitted from person-to-person Result in high mortality rates Cause public panic and social disruption Require special action for public health preparedness Bacillus anthracis Clostridium botulinum toxin Yersinia pestis Variola major (smallpox) Franciscella tularensis Ebola  CATEGORY B BIOTERRORISM AGENTS Second highest priority agents Moderately easy to disseminate Results in moderate morbidity rates and low mortality rates Require specific enhancements of CDC’s diagnostic capacity and enhanced disease surveillance Brucella species Epsilon toxin of Clostridium perfringens Food safety threats – Salmonella, E. coli O157:H7, Shigella Burkholderia mallei and pseudomallei Chlamydia Staphylococcal enterotoxin B Water safety threats – Vibrio cholerae RULE OUT AND REFER FLOWCHART