Long Exam 3: Spirochetes and Treponema PDF

LONG EXAM 3 Lesson 1: SPIROCHETES LEPTOSPIRA SPIROCHETES Members of phylum Spirochaetota Gram-negative Spiral/corkscrew shape Periplasmic flagella or endoflagella (axial filaments) Transverse fission Exhibit...

LONG EXAM 3 Lesson 1: SPIROCHETES LEPTOSPIRA SPIROCHETES Members of phylum Spirochaetota Gram-negative Spiral/corkscrew shape Periplasmic flagella or endoflagella (axial filaments) Transverse fission Exhibits spinning/twisting motility L. alexanderi L. alstoni L. borgpetersenii L. interrogans Tightly coiled, thin, flexible with bent ends that form a hook Gram-negative Stained with silver stain Best seen using a dark-field microscope It is very delicate, may appear only as a chain of minute cocci  Two axial endoflagella, anchored at It is the most common zoonotic opposite ends disease in the world Spinning motility Common in the tropics Obligate aerobe, oxidation of long-chain fatty acids Transmission Cannot use amino acids or Water or soil contaminated with the carbohydrates as major energy urine or blood of animals sources Usually rodents can come in contact Main source of nitrogen: ammonium with open wounds salts After hurricanes, the no. of cases Can survive for weeks in water, increase as animal urine can be particularly at alkaline pH carried by rain water Incubation is 1-2 weeks In the Philippines, common Leptospira found in rats are L. interrogans serovar Manilae, Losbanos, and Carlos Virulence Factors Heme-oxygenase - degrades heme, allowing Leptospira to obtain heme-iron OmpA-like Loa22 protein - present in pathogenic strains of Leptospira, absent in non pathogenic strains LPS, hemolysins and sphingomyelinases Clinical Features Two clinical syndromes Anicteric phase ○ Self-limiting ○ Non-specific flu-like symptoms ○ Sudden onset, rarely fatal ○ May recur after a few days (immune stage) Associated Disease Icteric phase Leptospirosis ○ Known as Weil’s Disease Disease caused by Leptsospira ○ Severe form w/c includes bacteria from contaminated soil or fever, renal failure, jaundice, water  respiratory distress and via darkfield microscopy hemorrhage A serological test, the gold standard ○ May last weeks or months if Patient’s sera is mixed w/ live the patient survives leptospires and observed for agglutination Detection/Diagnosis Indirect ELISA (IgM antibodies) Specimen: blood, CSF, urine Gram staining and silver staining may Better grown in liquid or semi solid be unreliable media Fletcher’s medium, Stuart’s medium, Treatment/Prevention EMJH Doxycycline and penicillin; (Ellenhousen-McCullough-Johnson-H doxycycline can also be prophylactic arris), Tween 80-albumin but dosage depends on risk Medium can be treated with 5-fluorouracil w/c kills non-leptospires Incubation: aerobic, 28 to 30 deg C, up to 8 weeks (slow generation time 6-12 hours) Dinger’s ring - diffuse zone of growth near the top of the tube consistent with the mcgs optimal oxygen tension TREPONEMA Detection/Diagnosis Microscopic agglutination test (MAT)  T. pallidum subsp. pallidum - syphilis Associated Disease T. pallidum subsp. pertenue - yaws Syphilis T. pallidum subsp. endemicum - bejel Systemic bacterial infection in T. carateum - pinta humans Due to its many manifestations, it is Treponema pallidum subsp. Pallidum named as the “great imitator and Gram-negative; thin spirochetes mimicker” Viewed using immunofluorescence, silver stain, darkfield microscopy Transmission With outer sheath or Sexual contact (STD) - oral, anal, glycosaminoglycan coating exterior vaginal to outer membrane Congenital transmission Endoflagella begin at each end and Contaminated needles wind around extending to and Blood overlapping at the midpoint It remains viable in whole blood or Generation time: 30 hours plasma stored at 4 deg C Microaerophilic; natural host is The mcg is sensitive to heat, cold humans and oxygen exposure Obligate intracellular pathogen; Infectious dose: 4 to 8 cells unculturable Virulence Factors Hyaluronidase - depolymerizes hyaluronic acid w/c creates microchannels Outer membrane proteins - promotes cell adherence; sparse in no. Fibronectin - protects the organism  from phagocytosis Clinical Features Primary Syphilis A single painless firm chancre at the point of infection w/c heals on its own (3 to 6 weeks) It is highly infectious Not treating primary syphilis at this stage will progress to secondary syphilis Secondary Syphilis Condyloma lata Pale, painless, cutaneous wart/growth Found in axillae, mouth or anogenital region Appear 2 to 4 weeks after primary syphilis Red maculopapular rash anywhere around the body Contagious secondary lesions that will subside (may recur after 3-5 years but not contagious) Systemic infection may occur (nephritis, meningitis, hepatitis, etc.)  Congenital Syphilis Miscarriages Tertiary Syphilis CNS anomalies Late symptomatic that can occur Saddlenose years or decades after initial infection Hutchinson’s triad: interstitial keratitis (20 to 30 years) (blind), sensorineural (deaf) and teeth Cardiovascular syphilis, neurosyphilis malformations (notch-pegged shape) (neurological symptoms), gummatous Bone deformities: saber shin, maxilla syphilis (formation of gummas or deformation or “bulldog-like lesions in bone, skin, liver) appearance” Exaggerated tissue response due to hypersensitivity to the mcg Can develop even if the patient did not experience primary or secondary syphilis Diagnostic/Detection Nontreponemal test To detect reagin from blood or CFS (presumptive) Reagin IgM and IgG autoantibodies produced in response to the lipids released during cell damage in syphilis infection Not exclusive to treponemes, other Diagnostic/Detection disease could also be detected Treponemal test E.g., leprosy, TB, etc. ○ To detect antibodies specific Antigen used: against TP antigen cardiolipin-cholesterol-lecithin (confirmatory) complex ○ Patient’s serum are tested TP-PA (T. pallidum-particle Rapid Plasma Reagin (RPR) Test agglutination) Toluidine Red Unheated Serum Test ○ Gelatin sensitized w/ TP (TRUST) antigen Unheated Serum Reagin (USR) Test TPHA (T. pallidum hemagglutination) Venereal Disease Research ○ Sheep erythrocytes Laboratory (VDRL) Test sensitized w/ TP antigen MHA-TP ○ Microhemagglutination Fluorescent treponemal absorption test (FTA-ABS) ○ TP cells (antigen) + patient serum (antibodies) + fluorescent conjugate = fluorescent spirochetes  Other T. Pallidum subspecies Bejel, pinta, and yaws Endemic treponematoses found in tropical, or dry and hot regions Nonvenereal but can be spread by body contact Diagnosis/Detection Microscopic detection of treponemes via darkfield, silver staining, immunofluorescence microscopy PCR test Specimen: tissue from lesions, CSF or blood Treatment: Penicillin G (most recommended), ceftriaxone, tetracycline or doxycycline Prevention: ○ Early detection and treatment ○ Proper use of condoms can reduce the risk of contracting disease  B. recurrentis B. burgdorferi B. afzelii B. garinii Larger than other spirochetes Microaerophilic Stains readily with bacteriologic dyes, Giemsa stain or Wright stain Multiple endoflagella (>7) without outer sheath Cultured in fluid media containing blood, serum, or tissue Repeated transfer in vitro can weaken their pathogenicity Can be passed from generation to generation (transovarial transfer) in BORRELIA ticks Antigenic structure can change during the course of infection ○ Antigenic variation is a virulence factor Linear chromosome  Diagnosis/Detection Culture in Barbour-Stoenner-Kelly (BSK II) medium supplied with rifampin, fosfomycin and amphotericin B (6 to 8 weeks incubation) Two-stage serology method Screening via enzyme immunoassay or indirect fluorescent antibody test Confirmatory via immunoblot/western blot Borrelia burgdorferi, B. afzelii, B. garinii Associated Disease Lyme Disease A zoonotic, vector-borne disease caused by several species of Borrelia Transmission Via Ixodes tick bite (ectoparasite of rodents, deer, domestic pets and even humans) Incidence rate peaks from May to August w/c is the peak of nymphs Most cases are associated with nymphal deer ticks Clinical Features Treatment Erythema migrans or “bull’s eye rash” Penicillin Late manifestation of flu-like Doxycycline symptoms Cefuroxime Arthralgia, arthritis, cardiac and Parenteral Ceftriaxone neurologic manifestations Prevention Endemic relapsing fever is Wear long-sleeved shirts and long transmitted via Ornithodoros soft pants when visiting tick infested ticks areas Zoonotic - rodents, small mammals, Use insect repellents soft ticks as reservoir E.g., DEET and permethrin Use light colored clothing to better see ticks that clings Avoid woody areas during peak months In PH, lyme disease is not common Clinical Manifestations Incubation period is 3-10 days Febrile: ○ Sudden onset of fever with chills ○ Abrupt rise in body temperature w/c lasts for 3-5 days ○ Bacteria is found in blood Afebrile stage: ○ Lasts for 4-10 days ○ Bacteria is not found in the blood Borrelia recurrentis and Borrelia spp. Recurrence of chills, fever, intense Associated Disease headache and malaise Relapsing fever Succeeding attacks become less ○ Vector-borne disease caused severe, could last up to 3-10 by Borrelia recurrentis recurrences (epidemic) and Borrelia spp. The changing antigenic variants may (endemic) allow immune system to select certain antigens only Transmission ○ Thus, relapse and lengthy Epidemic relapsing fever is infection occurs transmitted via human body louse In one patient, there could be many (Pediculus humanus) antigenic variants Humans are the only reservoir Diagnosis/Detection Lesson 2: ENTEROBACTERIACEAE Smears ○ Thin or thick blood smears stained with Wright or Giemsa ENTEROBACTERIACEAE ○ Look for large, loosely coiled spirochetes among the blood Classification cells Unlike the previous groupings where Animal inoculation we grouped together taxonomically ○ White mice or young rats are heterogeneous mcgs, this group falls inoculated intraperitoneally under the same family. with patient’s blood However, this family is very large, ○ After 2-4 days, tail blood is thus it is further subdivided into collected and examined for different “tribes” w/c is also divided spirochetes under different genera. Treatment Tetracyclines Erythromycin Penicillin Found in the mammalian intestines Defining Characteristics Ferments glucose/dextrose Reduces nitrates to nitrites Oxidase (-), except Plesiomonas Motile, except Klebsiella, Shigella, Yersinia Colony and Microscopic Morphology Non-sporeforming, gram (-) bacilli or coccobacilli Frequently used medium - MacConkey (MAC), Hektoen Enteric Agar (HEA) and Xylose Lysine Deoxycholate (XLD) Agar ○ HEA and XLD for Salmonella https://www.bbc.com/future/article/2023070 detection 6-the-troubling-rise-in-congenital-syphilis Facultative anaerobes  ○ Site of origin should be considered ○ Should be processed quickly but can be placed in transport media (Amies, Stuart or Cary-Blair) Has fast doubling time, may affect accuracy of quantity ○ Site of origin must be considered; sterile body sites where enterics are isolated signifies infection Virulence and Antigenic Factors Media for Isolation O-antigen - heat-stable antigen BAP and/or CAP + found in cell wall, stimulates early selective/differential media e.g., MAC antibody production MAC - LFs (bright pink) vs. NLFs H-antigen - heat-labile antigen found (clear or colorless) in flagella, stimulates late antibody ○ Lactose fermenters vs. production Non-lactose fermenter ○ Labile = heat sensitive HEA, XLD, MAC - highly selective K-antigen - heat-labile antigen found media for stool samples in capsule, stimulates late antibody Cefsulodin-Irgasan-Novobiocin (CIN) production - for selecting Yersinia Extended Spectrum Beta-Lactamases (ESBLs): inactivate extended Biochemical Identification spectrum antibiotics e.g., penicillins, Carbohydrate Fermentation cephalosporins and aztreonam ○ Ortho-Nitrophenyl-Beta-Galac ○ Beta-lactamases are enzymes toside (ONPG) - detects beta produced by the bacteria galactosidase against beta lactams ○ Triple Sugar Iron (TSI) - (antibiotics) fermentation of glucose, Fimbriae: attachment pili sucrose, lactose and H2S Aerobactin: iron chelating enzyme production (siderophore) ○ Methyl red and ○ Function as co-factor Voges-Proskauer (VP) - Cytotoxic-necrotizing factor 1 glucose metabolism Hemolysins Indole production - detects tryptophanase Diagnosis/Detection Citrate utilization - ability to use Collection and Handling:  citrate as C-source Urease production - hydrolysis of Escherichia coli urea to ammonia Consider as an opportunistic Lysine Iron Agar - ability to pathogen deaminate lysine ○ Most are harmless Two categories of Enterics Most significant species of the genus ○ Opportunistic pathogens - and a common bacteria of the colon commensal of the intestines Important potential pathogen of but can cause disease in humans other parts ○ Primary Intestinal pathogens Associated Diseases - not commensals, produce UTI and kidney infections: E. coli is infection when ingested e.g., the most common cause as it can Shigella, Yersinia and adhere to the epithelial cells of in the Salmonella urinary tract Ubiquitous in nature such as in soil, Septicemia and meningitis: E. coli is water and sewage the most common cause in neonates Commensal of the intestines of as it can be acquired in the birth mammals w/c associates them with canal fecal contamination “fecal Bacteremia caused by genitourinary coliforms”. tract infection or from the ○ Used as indicator for potable gastrointestinal tract water Stools contains many enteric Gastrointestinal Infections commensals, thus efficient screening ○ Enteropathogenic E. coli is needed to isolate pathogens. (EPEC) Enteric pathogens are generally Diarrhea in infants lactose-negative and children Pathogenic enterics are usually Outbreaks in Salmonella, Shigella, Aeromonas, nurseries Campylobacter, Yersinia, Vibrio, and Mucus in stool but no E. coli 0517:H7 blood Lactose fermenters: E. coli, ○ Enterotoxigenic E. coli (ETEC) Klebsiella, Enterobacter Traveler’s diarrhea Lactose fermenters w/ mucoid Watery diarrhea, no colony: Klebsiella or Enterobacter blood H2S production: Proteus, Salmonella, Caused by Citrobacter enterotoxins, Non-motile: Klebsiella or Shigella self-limiting VP (+): Klebsiella, Enterobacter or Lactose positive Serratia ○ Enteroinvasive E. coli (EIEC)  Dysentery w/ bowel endothelium penetration and the Invasion and glomerular destructive to the basement intestinal mucosa membrane Watery diarrhea with Hemorrhagic colitis, blood TTP (type of purpura), Lactose negative hemolytic-uremic ○ Enteroaggregative E. coli syndrome w/c results (EAEC) in low platelet count, Adheres to the hemolytic anemia, mucosal surface of kidney failure and the intestines death Watery diarrhea ○ Diffusely adherent E. coli (DAEC) Associated with UTI and diarrhea in children and pregnant women ○ Enterohemorrhagic E. coli (EHEC) More severe form EHEC O157:H7 Transmission: undercooked meats, unpasteurized milk and apple cider have spread the disease Shiga toxin Binds to the kidney endothelium EPEC prompting ○ Bf pilli inflammatory EHEC response of ○ Shiga toxin macrophages ETEC and ○ Heat stable & heat labile neutrophils EAEC w/c damage ○ Shigella toxin the EIEC  DAEC ○ Does NOT ferment sorbitol ○ Identified by serotyping (latex Diagnosis/Detection agglutination) Dry, pink, colonies on MAC Binding of the Black colonies with green metallic antibodies & antigen sheen in EMBA Ferments glucose, lactose, trehalose, Treatment and xylose Fluid management Indole (+) Mostly self-limiting but antibiotics can Methyl Red (+) be used Voges-Proskauer (-) ○ Except for Shiga toxin Citrate Utilization (-) producing EC Negative for H2S production Usually motile, but not all KESH Group Klebsiella, Enterobacter, Serratia and Hafnia Usually found in GI tract Associated with pneumonia, wound infections and UTI ○ Usually nosocomial Citrate positive H2S negative Phenylalanine deaminase negative MR negative VP positive Klebsiella spp. Found in GI tract Most commonly isolated Produces polysaccharide capsule which helps in immunoevasion and makes its colonies mucoidal ○ Yeast-like odor Enterohemorrhagic Escherichia coli O157:H7 Lactose (+) Cause severe intestinal infection in Urease (+) humans Non-motile Produce Shiga toxins Frequent cause of nosocomial Clinical Symptoms pneumonia ○ Starts with a watery diarrhea then progresses to bloody diarrhea ○ No WBCs are found in stool Laboratory Diagnosis mucoidal colonies (left) clear zone = capsule (right) Enterobacter spp. Comprised of 12 species: E. cloacae and E. aerogenes Isolated from wounds, urine, blood and CSF Major Characteristics capable of producing pigment when grown at room temp ○ Motile ○ MR (-) Hafnia spp. ○ VP (+) Only one species of Hafnia which is ○ SC (-) H. alvei Has been isolated (anatomical parts) E. cloacae and E. aerogenes from humans including stool Indole (-) Not normally pathogenic but may MR (-) cause nosocomial infections VP (+) especially in immunocompromised SC (-) patients Exhibits delayed Simmons Citrate Serratia positive reaction Comprised of 7 species: S. ○ Increase incubation period marcescens ○ Clinically signficant Proteus, Morganella and Providencia Nosocomial UTI and respiratory Normal intestinal microflora but can illness, bacteremia, septicemia, be opportunistic cardiac surgery, burn infections They are often referred to as Proteea Resistant to some antibiotics Phenylalanine deamination (+) Slow lactose fermenter and produce Lactose (-) pink colonies optimally at room temperature Proteus spp. Clinically significant species: P. vulgaris and P. mirabilis Can cause urine, ear, wound and  bacteremic infections meningitis and sepsis Strong urease (+) Ferments lactose, slow urea Phenylalanine deaminase (+) reaction hydrolysis Lactose (-) MR (+) SC (+) Resembles Salmonella spp. Salmonella spp. Primary intestinal pathogen One of the most well known clinically significant enterics Transmitted via fecal contamination of ingested food or water Its presence is a significant indicator of contamination Morganella ○ Thus, it is part of routine M. morganii is the only species under microbiological testing in the genus food, food products and Documented cause of UTI water Isolated from other anatomical sites Lactose (-) Urease positive VP (-) Phenylalanine deaminase positive Phenylalanine deaminase (-) Urease (-) Providencia Commonly produce H2S Two clinically significant species ○ Indicator for Salmonella is Providencia rettgeri is a pathogen of formation of black colonies urinary tract ○ Responsible for “rotten egg” ○ Has caused nosocomial odor outbreaks Inhibited by potassium cyanide Providencia stuartii can cause nosocomial outbreaks in burn units and has been isolated from urine Both are phenylalanine deaminase positive Citrobacter spp. Clinically significant species: C. freundii and C. koseri ○ Can cause nosocomial infections in the urinary tract, pneumonia, diarrhea, abscesses, and rarely  Clinical Manifestations: vomiting, chills, watery diarrhea, abdominal pain ○ Infection dose is quite high Treatment: self-limiting in immunocompetent individuals, antibiotics and antimotility medications may prolong symptoms (not recommended), fluid management ○ Antimotility medication modify normal peristaltic movement of gut Salmonella Virulence Factors Typhoid Fever Toxins Life threatening infection caused by Antigens S. enterica serotype typhi (S. typhi) Capsule Clinical Manifestations: prolonged Attachment appendages fever, chills, bacteremia, headache, ○ Fimbriae abdominal pain, nausea, vomiting Motility appendages Incubation: 9 to 14 days after ○ Flagella ingestion Reticuloendothelial (RE) system is involved - tissue/organs involved in the removal of dead or damaged cells ○ E.g., Liver, spleen, etc. Usually disseminate through various organs Common in tropical and subtropical regions Treatment: Antibiotic therapy, the type of antibiotic administered depends on where the infection was Associated Diseases picked up, ciprofloxacin is a common Acute gastroenteritis or food poisoning antibiotic used Transmission: improperly cooked food Typhoid Fever Pathogenesis ○ E.g., Eggs, chicken, milk, contaminated utensils, handling of pets Incubation: 8 to 36 hrs after ingestion  Shigella spp. Closely related to Escherichia Not a normal gut microflora Can cause bacillary dysentery (bloody, mucous, WBCs) Group A - S. dysenteriae Group B - S. flexneri Group C - S. boydii Group D - S. sonnei 2 to 5% of infected people will Lactose (-) become chronic carriers after Urea (-) infection H2S production (-) Mary Mallon “Typhoid Mary” Lysine decarboxylase (-) ○ Well documented case of a Citrate (-) typhoid carrier who worked No H-antigen as a cook ○ Lack of flagella Bacteria is spread through feces and Non-motile gallbladder is the reservoir site Endotoxin - LPS Gallbladder removal can solve the Exotoxin - Shiga toxin problem Humans are the only reservoir Low infective dose