Atypical Bacteria Part 1 PDF

Atypical Bacteria Part 1 B.A. Buxton, Ph.D.  Atypical Bacteria Intracellular Extracellular Chlamydia Legionella Mycobacterium Mycoplasma...

Atypical Bacteria Part 1 B.A. Buxton, Ph.D.  Atypical Bacteria Intracellular Extracellular Chlamydia Legionella Mycobacterium Mycoplasma Spirochetes Rickettsiales Leptospira Borellia Treponema Recommended Reading and Learning Objectives Recommended reading: appropriate chapters or sections from Medical Microbiology; Murray et al on line-in library With regard to each pathogen species describe: – Microbiological features- nature of cell wall, intra- or extracellular – where are they found in nature (are they strict human pathogens, or do they infect animals, or are they found in soil, etc) – how they are transmitted- ie, how do we become infected – the clinical manifestations of the diseases they cause, and be able to recognize a description of the disease in a test question – the pathogenesis of diseases they cause, being sure to relate bacterial virulence factors to the pathogenesis if discussed – Diagnostic features- how is an etiologic diagnosis made – how these infections can be prevented (ie, vaccine availability or behavioral modification, etc) Be able to define all terms in the ppt, including, but not limited to those in red. If you cannot do this from the lecture, you must look them up. Answer all questions posed in the ppt.  Bacteria Covered in This Lecture Chlamydia and Chlamydophila Legionella Mycobacterium  Case A 19-year-old male presented to an STD clinic with a purulent discharge from the urethra. A gram stain of the discharge is shown. A nucleic acid amplification test was ordered to determine the etiology. Questions answered in this section: – What is the most likely clinical diagnosis? – What is the most likely etiological diagnosis? – How did he become infected? – What is the pathogenesis of this infection? – What other kinds of infections does this group of organism cause?  Non-gonococcal Urethritis No gram-negative diplococci seen on gram stain of urethral discharge Microbial causes: – Chlamydia trachomatis – Ureaplasma urealyticum – Mycoplasma hominis – Mycoplasma genitalium – Trichomonas vaginalis  Chlamydia and Chlamydophila Common, worldwide human pathogens Although they do not gram stain, cellular characteristics similar to gram-negative – Outer membrane – LPS Obligate intracellular pathogens Cannot be cultured on agar or in broth- must be isolated in cell culture Once thought to be viruses (filterable) however they contain both DNA and RNA, ribosomes, make their own macromolecules Energy parasites- use host ATP  Chlamydia and Chlamydophila Unique life cycle found in no other bacterial group Exist in 2 forms- elementary and reticulate bodies – Elementary bodies- metabolically inactive, infectious form – Reticulate bodies- metabolically active, non-infectious form Life Cycle of Chlamydia- more info Note- a persistent infection of cells may occur in presence of gamma-IFN  Important Members of Group Chlamydia trachomatis – Causes ocular infections, genital infections, neonatal infections – Many serotypes- makes creation of vaccine more difficult – Enables repeat infections Chlamydophila pneumoniae – Respiratory infections Chlamydia psittaci – Bird pathogen; causes psittacosis in humans Flu-like illness with lower respiratory symptoms following exposure to bird excrement  Virulence Factors of Chlamydia Intracellular replication – Replicate in epithelial and other cells (not within macrophages) – Safe from complement, antibodies and phagocytes – Prevent fusion of endosome with lysosomes Endosome becomes inclusion body for replicating RBs Mechanisms for nutrient uptake from cytoplasm into inclusion body containing replicating RBs – Transporter proteins (Inc A etc) are inserted into membrane of inclusion body to form secretion systems – Secretion systems act as “molecular straw” taking up nutrients from cytoplasm; also allow for transport of bacterial molecules into host cell cytoplasm- ie proteins to inhibit apoptosis while RBs divide Role of secretion systems in survival and replication of Chlamydia Another look….  Pathogenesis of Chlamydia trachomatis Elementary bodies enter epithelial cells of conjunctiva and urogenital tracts Organism replicates as previously described Infected cells secrete inflammatory mediators (IL-1, IL-8, others), leading to acute, then chronic inflammation Disease manifestations due to chronic inflammatory response with tissue reorganization and scaring, especially if left untreated Infected cells stimulated with gamma-IFN can temporarily halt replication of, but do not kill RBs – May help lead to chronic infections with re-expression of disease at later date (see next slide for details) Persistently Infected Cells- more info Gamma-IFN- induces expression of cellular indoleamine-2,3-dioxygenase (IDO), which degrades tryptophan, thereby resulting in reduced levels of intracellular tryptophan.The lack of tryptophan leads to the death of Chlamydia spp. through tryptophan starvation. However, a population of Chlamydia spp. reticulate bodies (RBs) responds to the lack of tryptophan by acquiring a non-replicating but viable, persistent form. After removal of IFN- gamma and replenishment of tryptophan, the persistent forms of Chlamydia rapidly re-differentiate into infectious elementary bodies (EBs).  Chlamydia trachomatis Isolates exist as 15 serovariants that are subdivided into two major biovars: – Trachoma- caused by ocular tropic strains (A, B, Ba, and C) – Genitourinary tract tropic (D, E, F, G, H, I, J, and K) strains – Invasive lymphogranuloma venereum (L1, L2, and L3)  Disease Associations of Chlamydia trachomatis Trachoma – Ocular infection – Serotypes A-C STI- Genital discharge and perinatal infections – Urethritis, cervicitis, pelvic inflammatory disease, neonatal infections – Estimated to cause 4 million new infections in US/yr Most common bacterial STD in US – Individuals are frequently co-infected with N. gonorrhoeae – Serotypes D-K STI- Lymphogranuloma venerium – Serotypes L1, L2 and L3 – Uncommon in US  Chlamydia trachomatis Transmission/epidemiology – Human pathogen – no animal or environmental reservoirs – Transmitted by direct contact with infected secretions Trachoma- Spread from infected eye by contaminated fingers or flies Newborn conjunctivitis-Passage down birth canal of mother with genital infection Genital infections and lymphogranuloma venerium are transmitted by sexual contact  Trachoma Normal conjunctiva Caused by different serovars vs genital infections with of C. trachomatis Primarily a disease of people in developing countries (tropical and subtropical regions associated with crowding, poor sanitation, poverty) Transmitted from person to person on hands Begins as follicular conjunctivitis with diffuse inflammation Repeated infections lead to chronic inflammation of conjunctiva and eyelid; eyelid turns inward, eyelashes abrade cornea causing corneal ulceration, scarring, loss of vision Painful, debilitating 7 million people worldwide are blind due to trachoma Leading cause of preventable blindness in world Lymphoid follicles in infected conjunctiva  Trachoma Chlamydia eye infections Stain for elementary bodies  Urogenital infections Associated with C. trachomatis Begins as urethritis or cervicitis Both associated with purulent discharge If untreated, can progress – Males can develop prostatitis or epididymitis – Females can develop endometritis, pelvic inflammatory disease (salpingitis) – May lead to infertility  Pathogenesis of PID caused by C. trachomatis  Neonatal infections Associated with C. trachomatis Transmitted from infected mother to newborn at birth from organisms in vaginal canal Organisms contaminate infant’s eyes and nasopharynx leading to: – Ophthalmia neonatorum Incidence: 6.2/ 1000 live births in US Inflammation, hyperemia, purulent ocular discharge – Pneumonia  Chlamydophila pneumoniae – Clinical presentation Common infection in adults (200,000 – 300,000 cases/year) Most infections are asymptomatic Common in crowded conditions (schools, military barracks) Person – to – person transmission via inhalation of respiratory droplets Incubation: 3-4 weeks Clinical manifestations: Atypical (walking) pneumonia – Mild or asymptomatic – Often not associated with fever – Symptoms may resemble pharyngitis, bronchitis – Persistent non-productive cough – Most often involves a single lobe – 50 -75% of adults show serologic evidence of prior infection – Repeat infection is possible  Chlamydophila pneumoniae Common infection in adults Clinical manifestations: (200,000 – 300,000 cases/year) Atypical (walking) pneumonia – 50 -75% of adults show – Mild or asymptomatic serologic evidence of prior – Often not associated with infection fever – Repeat infection is possible – Symptoms may resemble Transmitted easily in crowded pharyngitis, bronchitis conditions (schools, military – Persistent non-productive barracks) cough Person – to – person transmission via inhalation of respiratory droplets Incubation: 3-4 weeks  Case A 54-year-old male renal transplant recipient presents to his physician with a 2 day history of malaise and myalgia and a one day history of fever (102F), SOB, non-productive cough, headache, abdominal pain and diarrhea. Abnormal breath sounds were heard on auscultation. Chest radiographs showed patchy alveolar infiltrates with consolidation in the lower right lobe. He was clinically diagnosed with pneumonia. Laboratory studies revealed elevated ESR (consistent with a significant inflammatory response), elevated liver function tests, hyponatremia and hypophosphatemia.  Case continued Induced sputum, BAL and blood was collected for culture and gram stain. No organisms were found on gram stain of BAL fluid or induced sputum. Two days after hospitalization, a urine antigen test was positive for Legionella pneumophila. After 5 days, routine cultures on nutrient agar and blood failed to grow any organisms, however growth was seen on buffered charcoal yeast extract (BCYE) agar. The patient’s empiric antibiotic was continued and the patient improved after several days of therapy.  Questions to answer: How did this patient become infected? What is the source of this organism? How does it cause disease? (ie, describe its pathogenesis) How can infections be prevented?  Legionella pneumophila Gram-negative cell wall structure, but does not pick up Gram stain well Coccobacilli in tissue; pleomorphic in culture Over 50 species with 70 This Gram-stained photomicrograph reveals chains, and solitary Gram- serogroups negative, Legionella pneumophila bacteria, which were found within a Legionella pneumophila sample taken from a victim of the serotypes 1 and 6 cause 1976 Legionnaires’ disease (LD) outbreak in Philadelphia. The over 90% of the human specimen was processed using the infections Gram-stain method, along with carbol fuchsin instead of safranin for the counterstain.  Legionella sp. Transmission/epidemiology – Aquatic saprophytes – Live in biofilms and within amoebae in the water – Found in wide variety of natural and man-made water sources- lakes, rivers, ponds, showers, fountains, spas, air conditioning units, room humidifiers, nebulizers – Transmitted in water mist- inhaled – Survive high temperatures and treatment with chlorine Hartmanella vermiformis filled with Legionella pneumophila Legionella in Man-Made Water Sources Legionella sp.- Associated Diseases Legionnaire’s disease/Legionellosis- pneumonia – A cause of both community-acquired and hospital-acquired pneumonia, especially in patients with risk factors- see below – ~ 10,000 reported cases of legionellosis/yr in US – Reportable disease – Infections more common in elderly and others with risk factors- smoking, renal or liver disease, diabetes, immune suppression due to cancer, AIDS, transplantation, corticosteriod use Pontiac Fever- mild self-limiting febrile illness – Fever, chills, myalgia, malaise  Pathogenesis of Legionnaires Disease (Legionella pneumonia) Infection through inhalation Phagocytosed by, and grows in alveolar macrophages Inhibits formation of phagolysosome, surrounding phagosome with materials from endoplasmic reticulum Incubation period of 2-10 days Abrupt onset of fever, chills, non-productive cough, headache Involves more than lung- multisystem disease involving GI tract (abdominal pain, diarrhea), liver(elevated LFTs) , kidneys (electrolyte disturbances), CNS (altered mental status) **No person-to-person transmission  Diagnosis of Legionellosis Clues to diagnosis: – Pneumonia with both pulmonary and non-pulmonary symptoms – Negative sputum gram stain (small very faintly staining, gm-negative bacilli, hard to see) PCR Urine antigen test Culture- best sample is Growth on Buffered bronchoalveolar lavage grown on Charcoal Yeast Extract Buffered Charcoal Yeast Extract (BCYE) agar agar (BCYE) Prevention of Legionella Infection Prevention efforts aimed at water sources If Legionella case/outbreak can be traced to source, treatments include: – Superheating of water to 70-80C – Install copper-silver ionization units Produce metallic ions that kill microbes – Hyperchlorination not recommended- organisms is tolerant of chlorine in water  A 32-year-old male presents with a 2 week history of fever, Case night sweats, weight loss and cough productive of blood-tinged sputum. The woman recently immigrated to the US from SE Asia and lives in a small inner city apartment with 7 family members including her father (71 years) and a 3 year old child. A chest radiograph reveals cavitary lesions in the upper lung lobes. Ziehl Nielsen staining of sputum reveals numerous acid-fast rods. A diagnosis of tuberculosis Tuberculosis  Global Impact of Tuberculosis One of leading causes of death globally Estimated 1/3 of world population is infected Estimated 8 million new cases yearly 1.5-3.0 million people die annually One of most common cause of death from infection globally Antibiotic resistance is major problem hampering control efforts  Mycobacterium Acid-fast, aerobic rods Cell wall contains lipids (mycolic acid) Waxy coat makes organisms resistant to drying and chemicals – Important for survival in and out of body Does not gram stain, despite peptidoglycan layer Many species- 3 of importance in humans: – M. tuberculosis- TB – M. leprae- leprosy – M. intracellulare- disseminated disease in immune suppressed  Mycobacterium Many species- 3 of importance in humans: – M. tuberculosis- TB-most important member of group – M. leprae- leprosy pockets of high endemicity still remain in some areas of Africa, South America and India – M. intracellulare- pneumonia; disseminated disease in immune suppressed TB incidence per 100,000 population Persons at Risk for TB in the US Foreign-born persons from areas where TB is endemic (SE Asia, Africa- see map next slide) Close contacts of person known or suspected to have TB Residents and employees of high-risk congregate settings (ie. Homeless shelters, prisons) People with HIV/AIDS Injection drug users Medically underserved, low-income populations  Pathogenesis of Mycobacterium tuberculosis Infection by inhalation of droplet nuclei Bacteria gain access to lung, replicate within alveolar macrophages Replication is very slow Outcome dependent on strength of host immune response – Latent infection – Active infection Pulmonary Non-pulmonary, disseminated involving multiple tissues/organs Overview of Pathogenesis  Latent Infections with Mycobacterium tuberculosis Immune competent – develop latent infection -bacteria are contained by strong cell-mediated response with formation of granulomas 90% of initial infections result in latent infection Over time, granulomas may become calcified nodules (Gohn complex) Bacteria within granulomas are unable to replicate but remain viable for years-decades  Active Infection Active disease can result upon initial infection (=primary TB) or following reactivation of latent infections (= Miliary secondary TB) TB Primary tuberculosis – Majority develop miliary TB: Progressive, disseminated disease (usually immune compromised, infants or children) – Pulmonary TB only occurs in about 6% Secondary (reactivated) tuberculosis – Immune suppression allows latent TB to reactivate – Granulomas break down releasing organisms – Pulmonary TB results – Most cases of active TB result from reactivated infections Pulmonary TB  Primary vs Reactivated Pulmonary TB Pulmonary disease can follow primary infection (~ rare) or reactivation (most common) Primary infection resulting in pulmonary disease occurs more commonly in children vs adults Reactivated disease most often follows infection of immune competent adult whose immune system is subsequently compromised  Infection with M. tuberculosis Pulmonary disease resulting Miliary TB from: Reactivated Involves Primary infection infection multiple organs Children Adults  Manifestations of Reactivated Pulmonary TB Insidious onset of fever, night sweats, weight loss, anorexia, malaise, weakness Cough often productive of blood-streaked sputum Apical and posterior segments of upper lobes most often affected In reactivated pulmonary disease, may see cavitary lesions on CXR  Miliary (Non-Pulmonary) TB Seen in children with primary infection and increasingly in TB patients co-infected with HIV Infected macrophages can take organism hematogenously throughout body Multiple organs can be affected: – Lymphadenitis – Meningitis – Pott Disease- involvement of the spine. Progressive back pain, can lead to paralysis if untreated – Chronic arthritis – Genitourinary involvement- flank pain, dysuria – Gastrointestinal- can involve any site in GIT with painful ulcers, malabsorption, diarrhea  Diagnosis of Patients with Suspected Active TB TB skin test and Gamma-IFN release assay – See next slide Acid fast stain of sputum – 45-80% sensitivity Sputum sample for nucleic acid detection, culture and sensitivity testing – NA detection using RNA probes and/or PCR can detect presence of Acid-fast stain of sputum M. tuberculosis Results in 1-2 days; 95% if smear-positive Culture is gold standard and must be done to assess antibiotic sensitivity – Culture can take 2-6 weeks CXR – Look for cavitary lesions Colonies grown on agar  Diagnosis of TB continued TB skin test – Purified protein derivative (PPD) injected intradermally – Within 48 hours, a positive lesion is erythematous and indurated – The area of induration is measured and interpreted – Can detect latent or active infection Interferon gamma release assay – Collect blood and separate T cells – Incubate T cells in presence of TB antigens or controls – Assay for production of gamma IFN – More consistent results compared to skin test – Can detect latent or active infection  Treatment of TB Use directly observed therapy to assure compliance- true globally! Treat latent TB for 9 months with 2-drug regimen Treat active TB as follows: – Isolates with no identified resistance use 4-drug regimen for 2 months, followed by 4-7 months of 2 drug regimen Resistance to anti-TB drugs is ~common- with multidrug and extensively drug resistant strains in some areas of world. Drug resistance is very serious issue!  Case A 23 year old AIDS patient had fever, night sweats, fatigue and diarrhea for the past 3 months. Over that time, she has lost 22 pounds and is short of breath. Her CD4 count has been below 50/cmm for the past 6 months and her viral load is 55,000/cmm. Routine blood cultures were negative after 48 hours; however, growth was seen after 10 days. DNA probes identified the organism as belonging to the Mycobacterium avium complex  Pathogenesis of Disseminated MAC In AIDS Patients Low power High power of same tissue Note acid fast rods in tissue Organisms disseminate throughout body Proliferate to high numbers in ~every organ Organ function disrupted  Questions What are the diagnostic features of this organism? What are the sources of this organism? What other diseases does it cause? How is it treated?  Mycobacterium avium complex (MAC) Group of mycobacterium that cause similar diseases Includes M. avium and M avium intracellularie Ubiquitous in soil wand water, infect multiple species of birds and mammals with or without causing disease Causes serious disseminated disease in immune compromised people (AIDS, cancer or transplant patients, others  Pathogenesis of Disseminated MAC Infection Disseminated MAC occurs only in severely immune compromised (CD4 cell count below 50/uL) Enters body via ingestion or inhalation Crosses mucosal epithelium and infects the resting macrophages Infected macrophages carry organism throughout body including lymph nodes, liver, spleen, bone marrow, and other sites. Pathogenesis of cytokine storm Organism replicates to very high numbers within macrophages in various tissues Infected macrophages secrete cytokines – “cytokine storm” results with results shown in diagram