Non-Spore-Forming Gram-Positive Rods: Listeria & Gardnerella - PDF

Non-Spore-Forming Gram-Positive Rods Listeria monocytogenes Gardnerella vaginalis Dr. Nada A. Abdelrahim 1 Listeria monocytogenes 2 Disease Spectrum L. monocytogenes causes Meningitis and Sepsis in newborns, pregnant women, and immunosuppressed adults It al...

Non-Spore-Forming Gram-Positive Rods Listeria monocytogenes Gardnerella vaginalis Dr. Nada A. Abdelrahim 1 Listeria monocytogenes 2 Disease Spectrum L. monocytogenes causes Meningitis and Sepsis in newborns, pregnant women, and immunosuppressed adults It also causes outbreaks of Febrile Gastroenteritis It is a major cause of concern for the food industry 3 General Characteristics L. monocytogenes is Small Gram-Positive Rod arranged in V- or L-shaped formations similar to corynebacteria It has an unusual Tumbling Movement that distinguishes it from corynebacteria (which are nonmotile) Colonies on Blood Agar plate produce a narrow zone of β-hemolysis that resembles the hemolysis of some streptococci. Listeria grows well at cold temperatures… so, storage of contaminated food in refrigerator can increase risk of gastroenteritis The ability of growth in cold temperature is called “Cold Enhancement" 4 5 6  Although L. monocytogenes is actively motile by means of Peritrichous Flagella at room temperature (20 − 25 °C), the organism does not synthesize flagella at body temperatures (37 °C): ❖ At 28oC it is Motile (1-5 peritrichous flagella) ❖ At 37oC it is less to No Motility ❖ On liquid medium ------ "tumbling motility" (see video) ❖ On solid medium ------- "umbrella growth“ (see next slide) CAMP Test is used for the Identification of Listeria monocytogenes (see next slide) 7 Umbrella motility of Listeria monocytogenes grown at room temperature in semi-solid agar medium 8 CAMP Test for the Identification of Listeria monocytogenes CAMP-positive Listeria monocytogenes inoculated at right angles to beta-hemolytic Staphylococcus aureus. Note the arrow-shaped zone of weak enhanced hemolysis indicating a positive CAMP test 9 10 11 Lactose fermenting colonies on MacConkey agar medium 12 Pathogenesis Listeria infections occur primarily in 2 Clinical Settings: (1) in Fetus or in Newborn (as a result of transmission across the placenta or during delivery) (2) in Pregnant Women and Immunosuppressed Adults (especially renal transplant patients) * Note that pregnant women have reduced cell-mediated immunity during the third trimester 13  Listeriae is distributed worldwide in animals, plants, and soil It is transmitted to humans primarily by ingestion of unpasteurized milk products, undercooked meat, raw vegetables, and contact with domestic farm animals and their feces Listeriosis is primarily a foodborne disease associated with eating unpasteurized cheese and delicatessen meats in developed countries (e.g., US) 14  Following ingestion, the bacteria appear in the colon and then can colonize female genital tract From this location, they can: infect the fetus if membranes rupture, or infect the neonate during passage through birth canal Pathogenesis of Listeria depends on the organism's ability to invade and survive within cells Invasion of cells is mediated by internalin made by Listeria and E-cadherin on the surface of human cells The ability of Listeria to pass the placenta, enter the meninges, and invade the gastrointestinal tract depends on the interaction of internalin and E- cadherin on those tissues 15  Upon entering the cell, the organism produces listeriolysin (allows it to escape from phagosome into the cytoplasm -thereby escaping destruction in phagosome) Because Listeria preferentially grows intracellularly, cell-mediated immunity is a more important host defense than humoral immunity Suppression of cell-mediated immunity predisposes to Listeria infections L. monocytogenes can move from cell to cell by means of Actin Rockets/ Comet Tails (filaments of actin polymers) and propel bacteria through membrane of one human cell and into another (see next slide) Listeria can also move using flagella at temperatures of 30°C and below. But at 37°C, Listeria moves primarily using actin rockets 16 (A) Schematic diagram showing intracellular life cycle of L. monocytogenes. (B, C) Electron micrographs showing entry of L. monocytogenes into PtK2 cells and an actin comet tail isolated from PtK2 17cells Clinical Findings Infection During Pregnancy can cause: ❖ Abortion, ❖ Premature delivery, or ❖ Sepsis during the peripartum period Newborns Infected at the Time of Delivery can have: ❖ Acute Meningitis 1 to 4 weeks later Bacteria reach the meninges via the bloodstream: - Bacteremia Infected mother are either Asymptomatic or has an Influenza-Like Illness 18  L. monocytogenes infections in immunocompromised adults can be either Sepsis or Meningitis Gastroenteritis caused by L. monocytogenes is characterized by: ❖ Watery Diarrhea, Fever, Headache, Myalgias, and Abdominal Cramps But little Vomiting Outbreaks are usually caused by contaminated dairy products, but undercooked meats such as chicken and hot dogs and ready-to-eat foods such as coleslaw have also been involved 19 Laboratory Diagnosis Laboratory diagnosis is made Primarily by Gram stain and Culture Appearance of: - Gram-Positive Rods resembling diphtheroids and - the formation of Small, Gray Colonies with a Narrow Zone of β- hemolysis on Blood Agar plate Suggest the presence of Listeria Isolation of Listeria is Confirmed by: the Presence of Motile organisms (differentiate them from non-motile corynebacteria) Identification of the organism as L. monocytogenes is made by Sugar Fermentation Tests (L. monocytogenes ferments glucose) 20  Treatment Treatment of invasive disease (such as meningitis and sepsis) consists of: ❖ Ampicillin with or without Gentamicin ❖ Trimethoprim-sulfamethoxazole (can also be used) Resistant strains are Rare Listeria Gastroenteritis typically does not require treatment 21  Prevention Prevention is difficult because there is no immunization Limiting the exposure of pregnant women and immunosuppressed patients to potential sources such as farm animals, unpasteurized milk products, and raw vegetables is recommended Trimethoprim-sulfamethoxazole given to immunocompromised patients to prevent Pneumocystis pneumonia can also prevent listeriosis 22 Gardnerella vaginalis 23 Disease Spectrum Gardnerella vaginalis is the main organism associated with Bacterial Vaginosis This disease is the most common vaginal infection of sexually active women 24  General Characteristics Gardnerella vaginalis is a Small, Facultative Gram-variable Rod The term “Gram-variable" refers to the observation that some organisms are Purple while others are Pink in a Gram-stained specimen Structurally, it has a Gram-Positive Cell Wall. But the wall is thin, and older organisms tend to lose the purple color (see next slide) 25 26  Pathogenesis The pathogenesis of bacterial vaginosis is uncertain G. vaginalis is often found in association with Anaerobes (such as Mobiluncus and Prevotella) and Non-Anaerobes (such as Mycoplasma hominis and Ureaplasma urealyticum) Together they cause the symptoms of Bacterial Vaginosis Bacterial vaginosis is not thought to be transmitted by sexual activity, it is considered to be a dysbiosis in which the Lactobacillus that are found as normal flora in the vagina are replaced by these other organisms 27  Clinical Findings Bacterial vaginosis is characterized by a malodorous, white or gray-colored vaginal discharge The discharge has a characteristic "fishy" odor Inflammatory changes are typically absent, which is why it is called a "Vaginosis" rather than a “Vaginitis" Mild itching may occur Women with bacterial vaginosis have a higher incidence of preterm deliveries, and consequently, a higher incidence of morbidity and mortality occurs in their newborn children 28  Laboratory Diagnosis Clue Cells (vaginal epithelial cells covered with bacteria) are an important laboratory finding seen in a microscopic examination of vaginal discharge (see next slide) In addition, the “Whiff Test" (consists of treating the vaginal discharge with 10% KOH and smelling a pungent, "fishy" odor) is often positive However, trichomoniasis, which can also cause a positive whiff test, must be ruled out before a diagnosis of bacterial vaginosis can be made A pH of greater than 4.5 of vaginal discharge supports the diagnosis of bacterial vaginosis 29 Treatment and Prevention Drug of choice is Metronidazole Treatment of sexual partners is not recommended as it is not considered to be transmissible There is no vaccine 30 Clue cells In bacterial vaginosis - The lower epithelial cell is a ‘”clue cell’’ because its surface is covered with bacteria - The upper epithelial cell is not a "clue cell’’ because its surface has few bacteria 31 32 Wet mount appearance of clue 33 cells 34