Microbiology Oral Topics PDF

Microbiology oral topics Microbiology oral topics Viral clas 1. Clinically important viral inf...

Microbiology oral topics Microbiology oral topics Viral clas 1. Clinically important viral infections (beyond HIV and hepatitis) Lecture IX: viruses, GENERAL VIROLOGY Classification: Definition of virus and viral classifications: 1. Structure Virus – is a submicroscopic infectious agent that replicates only intracellularly in a living host 2. Biochemical properties o Viruses infect all types of life forms (animals, plants, bacteria and archaea); bacteriophages infect bacteria o They have no mechanisms to replicate themselves (e.g., enzymes, energetic pathways for replication without host cells) 3. Geographical localization o Their genetic information is enveloped within a protein coat 4. Diseases 5. … Virion structure: Virion structure Ebolavirus is an example for enveloped viruses Non-enveloped viruses are more stable against certain chemical and physical conditions than enveloped ones Virion structure www.biomedi o Enveloped viruses can be lysed easier because on their membrane proteins are receptors for attachment to the host cell and aked capsid virus (non-enveloped) phospholipids/glycolipids are more sensitive to certain chemical/physical conditions Enveloped viruses Naked capsid virus (non-enveloped): Enveloped virus with membrane and glycoproteins: Nucleic acid Membrane and glycoprote Enzymes, other proteins Structural proteins www.biomedic-plzen.cz Nucleocapsid Viral replication Virion structure: Viral replication: Virion structure Virion – virus outside of the host www.biomedic-plzen.cz DNA viruses Early Burst Late cancentration size Enveloped Non-enveloped Log virus RNA viruses Viral replication Time Infection Eclipse Latent (+RNA) (-RNA) (+/- RNA) (+RNA via DNA) Production Phases of viral replication: www.biomedic-plzen.cz www.biomedic-plzen.cz 1) Recognition and attachment to the target cell via receptor 2) Penetration into target cell 3) Uncoating (in cytoplasm or nucleus) Intracellular 4) viral reproduction Macromolecular synthesis (protein synthesis) 5) Release of viral particles 1 www.biomedic-plzen.cz Mur Microbiology oral topics Protein synthesis: Viruses use the components of the host cells: ribosomes, tRNA, mechanisms for post-translational modification In some RNA viruses, RNA is translated to a single big (macro)protein that is cleaved by proteases into functional smaller ones o E.g., in HIV, which can be used as a target for antiviral drugs SPECIAL VIROLOGY Papillomaviruses: Classification: dsDNA, nonenveloped, at least 100 types Pathogenesis: Infection and replication in squamous epithelium of skin and mucous membranes (genital, oral, conjunctival papilloma) Viruses: HPV (Human Papillomaviruses) Disease: dysplasia which can lead to cancer Prevention: vaccination Epidemiology: Papillomaviruses - Epidemiology Low risk HPVs are not important concerning cancer development (they are only responsible for e.g., STDs) High risk HPVs are responsible for dysplasia and the possible development of cancer In men with penis cancer, HPV is a negative factor in the outcome of the cancer à surgery must be done more extensive Papillomavirus Inoculation of epithelium Direct contact Sexual contact Hand, foot, thorat, cervix Prevention: Polyomaviruses: vaccination Classification: dsDNA, nonenveloped, at least 100 types Pathogenesis: causes asymptomatic infections o In immunocompromised patients, it causes progressive multifocal leukoencephalopathy (PML) and renal diseases (in patients after kidney transplantation) www.biomedic-plzen.cz Polyomaviruses - JC Viruses: BK (renal disease), Epidemiology (PML) Epidemiology: aerosol Virus Multiplication viremia in respiratory tract viremia Multiplication in kidney Reactivation Viruria, CNS Hemorrhagic cystitis Adenovirus: www.biomedic-plzen.cz Classification: linear, dsDNA, nonenveloped (can easily spread by airways) Pathogenesis: causes infections of mucoepithelial cells in the respiratory and GI tract, conjunctiva or cornea Prevention: vaccination for serotype 4,7 (mainly used for soldiers) Diseases: o Acute respiratory disease o Laryngitis, croup (=laryngotracheobronchitis; respiratory infection caused by virus, leading to swelling inside trachea), bronchiolitis o Conjunctivitis, epidemic keraconjunctivitis o Gastroenteritis + diarrhea Epidemiology: Virus is spread by direct contact via respiratory droplets, feces, direct contact via hands, inadequately chlorinated swimming pools Main affected groups: children < 14 years and people in crowded areas 2 Microbiology oral topics Herpesviruses: Classification: dsDNA, enveloped Detection: PCR (NO RT-PCT, because it is dsDNA) Herpesviruses and their properties: Spreads into Affects also Transmission Human herpesvirus 1 (HHV-1/HSV) Mucoepithelial cells Neuron Close contact Human herpesvirus 2 (HHV-2) Mucoepithelial cells Neuron STD Varicella-Zoster virus (VZV) Mucoepithelial cells Neuron Respiratory system and contact (HHV-3) Epstein-Barr virus (EBV) Epithelial cells B-cells Saliva (kissing disease) (HHV-4) Cytomegalovirus (CMV) Epithelial cells Monocytes, lymphocytes Lymphocyte tissue transplantation, (HHV-5) transfusion, congenital spread, contact A) HHV-1 and HHV-2 Causes significant morbidity and mortality on infections of the eye, brain or disseminated infections (especially in immunocompromised HIV patients and neonates) Treatment: Acyclovir (most-prescribed drug), Valacyclovir, Penciclovir, Famciclovir It can cause: Herpes pharyngitis Eczema herpeticum Herpetic keratitis Genital herpes (one of most common STDs) Herpetic whitlow (fingers) Herpes encephalitis (usually HHV-1) and meningitis Herpes gladiatorum (infection of the body) o Causes damage of neuronal tissues with long-term symptoms B) Varicella-Zoster Virus (VZV) It is causative agent of chickenpox (varicella) o Classical childhood exanthems (widespread rash occurring on the outside of the body and usually occurring in children) o Primary infection is usually more severe in adults (20-30% develop interstitial pneumonia) o Prevention: vaccination (but after normal infection, children have long-life immunity and common course of disease is not critical) It causes even in case of chickenpox immunity herpes zoster/shingles (viral infection causing painful rash, most common with a single stripe of blisters that wraps around either the left/right side of torso) o Postherpetic neuralgia can develop (affects nerve fibers and skin, causing burning pain that lasts long after the rash and blisters of shingles disappear Possible treatment (see CMV): Ganciclovir, Valganciclovir, Cidofovir, Foscarnet C) Epstein-Barr Virus (EBV) Causes heterophile antibody-positive infectious mononucleosis (glandular fever): o Symptoms: § Lymphadenopathy (have abnormal size and consistency) § (Hepato)Splenomegaly § Exudative pharyngitis (cannot be distinguished by physical examination from Streptococcus pyrogenes infection) Antibiotic therapy in case of EBV and not S. pyogenes can cause toxic side effects § High fever, malaise o Infectious mononucleosis can lead to chronic diseases, EBV-induced lymphoproliferative diseases or hairy oral leukoplakia D) Cytomegalovirus (CMV) It is a typical congenital infection (causes thrombocytopenia, microcephaly, rash) which can cause loss of hearing (after many years) à cytomegalic inclusion disease It can be also gained by perinatal infection or sexual transmission (STD) Infection of immunocompromised host causes: pneumonia, retinitis, colitis, esophagitis, failure of many kidney transplants (screening after transplantation necessary) Treatment: Ganciclovir, Valganciclovir, Cidofovir, Foscarnet (they all can also be used in VZV therapy) 3  Poxviruses - Smallpox Microbiology oral topics Variola virus: Is member of the PROXIVIRUSES Classification: complex large DNA viruses Diseases: smallpox (Orthopoxivirus) and molluscum contagiosum (Molluscipoxivirus) o Smallpox was totally eradicated in 1980, because it does not mutate and has no animal reservoir Pathogenesis: it first replicates in URT, followed by dissemination to internal and dermal tissues www.biomedic-plzen.cz Picornaviruses: Classification: small RNA viruses with a naked capsid Viruses: Enteroviruses, Rhinoviruses, Hepatitis A virus A) Enteroviruses Examples: Polioviruses, Coxsackie viruses, Echoviruses, Enteroviruses Pathogenesis: transmitted by fecal-oral route à URT à oropharynx à intestinal tract is the portal of entry into body à spreads by viremia à attacks different cells o Viremia – viruses enter the bloodstream and have access to the rest of the body Diseases: meningitis, encephalitis (both important in kids), rash, myocarditis, viral infect of the URT, paralytic disease (poliovirus; typical in children) Prevention by 2 types of vaccine: o Inactivated polio vaccine (IPV); safer than OPV o Lave attenuated oral polio vaccine (OPV) o Poliovirus was eradicated by vaccination (virus has no animal reservoir) B) Rhinoviruses The most important cause of common cold Paramyxoviruses: Classification: negative sense ssRNA, enveloped virus Examples: Measles virus, parainfluenza virus, Mumps virus, Respiratory syncytial virus, Metapneumovirus, Nipah virus, Hendra virus (leads to severe encephalitis) Pathogenesis: direct infection of neurons which can cause an immune mediated encephalitis A) Measles virus Measles symptoms: o Characteristic maculopapular rash (flat, red area on the skin that is covered with small confluent bumps) o Cough o Conjunctivitis, photophobia o Complications with otitis media o Bronchopneumonia o Purpura (red/purple discolored spots on the skin that do not blanch on applying pressure) o Petechiae (red/brown/purple spots on skin; small and do not change color when pressing; blood vessels bleed into skin) § Petechia is smaller than 2 mm, while purpura is larger Atypical measles: more intense rash, petechiae, purpura Subacute sclerosing panencephalitis: CNS manifestation B) Parainfluenza viruses Pathogenesis: infection of epithelial cells of URT; ubiquitous virus (=found everywhere) Clinical manifestation: mild cold-like upper URI, bronchitis and pneumonia Influenza viruses: Are member of the ORTHOMYXOVIRUSES Classification: negative sense RNA, enveloped virus Examples: Influenza A, B and C virus (A+B are most common) Pathogenesis: o It establishes a URT infection and kills mucous secreting and ciliated cells o In case of infestation of lower respiratory tract, the virus can cause serious desquamation of bronchial and alveolar epithelium o Low level viremia and virus can rarely invade other organs than lungs 4  Microbiology oral topics Replication and structure of the virus: Envelope structure contains hemagglutinin (HA, H) + neuraminidase (NA, N) + matrix membrane proteins Internal structure contains 8 different helical nucleocapsid segments + nucleoproteins + reverse transcriptase (it is a RNA virus) Replication occurs in the NUCLEUS Receptors: A) Hemagglutinin Viral attachment protein (designation: H1, H2, H3, H4, H5, etc.) Promotes fusion of the envelope to the host cell membrane B) Neuraminidase Glycoprotein which cleaves sialic acid of the cell membrane on glycoproteins Orthomyxoviruses – Influenza It prevents clumping and Viruses - Epidemiology facilitates the release of virus from infected cells Targe for two antiviral drugs: Oseltamivir (Tamiflu) and Zanamivir (Relenza) à therapy must be quick, otherwise it has no impact on the outcome Epidemiology/evolution of influenza virus: Human: Duck: H3N2 D1-D8 H1-H8 Pig – coinfection H3N2 D1-D8 H1-H8 Reassortment Clinical symptoms: www.biomedic-plzen.cz Fever, malaise, myalgia (muscle pain), sore throat, nonproductive cough, GI symptoms (abdominal pain, vomiting), otitis media Complications: viral pneumonia, bacterial pneumonia, encephalitis (inflammation of brain) Rhabdoviruses: Classification: negative sense RNA, enveloped virus Examples: Rabies virus Disease: Rabies virus infection; cannot spread via air, vaccination after bite is possible Pathogenesis: 1) Biting by rabid positive animal (e.g., bat, rat, mice, dog) 2) Replication in the muscles (minimum symptoms) 3) Infection of peripheral nerves 4) Infection of the brain 5) Death Filoviruses: Coronavirus: Classification: negative sense RNA, enveloped virus Classification: RNA, enveloped; there are different types of coronaviruses Examples: Marburg virus, Ebola virus Symptoms/Causes: causing respiratory tract infections (common cold 10-15%) Diseases: severe or fatal hemorrhagic fevers Severe acute respiratory syndrome-related virus (SARS) o First seen in 2003/2004 Rotaviruses: o High mortality in elderly Member of REOVIRUSES o Incubation period of 2-14 days, fever > 38°C, non-productive caught and hypoxia Classification: dsRNA, non-enveloped virus Middle East respiratory syndrome-related virus (MERS) Disease: infantile diarrhea (can lead to severe dehydration) o First seen in Saudi Arabia in 2012 o Also identified in camels and bats Pathogenesis: virus replicates in epithelial cells of the small intestine o Causes respiratory distress syndrome with mortality rate of 40% Prevention: vaccination of newborns 1-2 month after birth Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Causes coronavirus disease 2019 (COVID-19) First seen in Wuhan in 2019 Infects the host via ACE2 receptor (humans and animals) Disease: fever > 38°C, myalgia, non-productive caught, pneumonia, hypoxia Laboratory diagnostics: RT-PCR (via nasopharyngeal swab), antigen detection (direct), antibody detection (indirect) 5 Microbiology oral topics Togaviruses and Flaviviruses: Classification: positive sense RNA Examples: Arboviruses, Rubella viruses, Hepatitis C virus Diseases: o Zoonotic diseases – vectors are mosquitos or ticks o Arboviruses: Eastern equine encephalitis, mild systemic disease o Flaviviruses: encephalitis, fever, Dengue fever, Yellow fever, West-Nile fever Rubella virus: Disease: rubella Pathogenesis: classic childhood exanthems (widespread rash occurring on the outside of the body and usually occurring in children) o It has the same symptoms as measles o Can cause severe congenital defects until the 20th week of pregnancy (ocular defects, heat defects, deafness, microencephaly, mental retardation) § It can lead to abortion Prevention: vaccination Bunyaviruses and Arenaviruses: Classification: negative sense RNA, enveloped Examples: Bunyavirus, Phlebovirus, Hantavirus Diseases: usually zoonotic diseases – vectors are mosquitos, ticks or rats Symptoms: o Retroviridae Hemorrhagic fever (affect many organs, damage blood vessels and the affect the body’s ability to regulate itself) § Caused by Marburg virus, Ebola virus, Lassa fever virus and Yellow fever virus o Encephalitis Human immunodeficiency virus (HIV) o Conjunctivitis o Febrile illness Destruction of CD4+ T-cells o Respiratory distress syndrome (when fluid builds up in alveoli; less O2 reaches the bloodstream) Retroviruses: Binding of HIV to CD4 receptor Classification: two RNA molecules, enveloped Properties: After the decrease bellow 200/µl opp o Contain reverse transcriptase (RNA à DNA; RNA dependent DNA polymerase) o appeared - AIDS They integrate into the genome of the host cell (treatment is difficult) Examples: Alpha-/Beta-/Gamma-/Deltaretrovirus, etc. Primate T-lymphotropic virus (PTLV-1-3): Example: Deltaretrovirus o Retroviridae It leads to human TLV (HTLV) or simian TLV (STLV) à induction of leukemia and lymphomas from T-cells 2. HIV and hepatitis Human immunodeficiency virus (HIV) Human immunodeficiency virus (HIV): Structure: Types: HIV-2 (prevalent in Africa and evolutionary older) and HIV-1 (firstly occurred in late 1970s/early 1980s) Two identical copies of +RNA (mRNA) HIV leads to AIDS (acquired immunodeficiency syndrome) Structure: 10 – 50 copies of reverse transcriptase Two identical copies of +RNA (mRNA) Integrase enzyme Two cellular tRNA (transfer RNA) 10-50 copies of reverse transcriptase and integrase enzymes www.biomedic-plzen.cz Two cellular tRNA (transfer RNA) Specialty of HIV virus: They have affinity to CD4+ cells and bind to their receptor à leads to destruction of CD4+ cells After the decrease below 200 CD4+ cells/!L, opportunistic infection appears, leading to AIDS o With a CD4 count below 200, the body is vulnerable to opportunistic infections o These are infections the immune system can usually prevent on its own, but with low CD4 count, the immune system is not able to fight them off o CD4 is a glycoprotein found on TH cells, monocytes, macrophages and dendritic cells 6 www.biomedic-plzen.cz Microbiology oral topics Spread: Sexual contact Sperm Body fluids (especially blood) Vaginal secretions No spread: kissing (if no injury is present), insect bite There is a higher risk from positive man to woman o Semen contains high concentration of virus with a bigger contact surface of mucous membrane in the vagina § 1/100 – 1/1000 in vaginal sex/intercourse à positive man to negative woman § 1/10 in anal sex/intercourse o From positive woman to negative man, the probability is way lower, as the virus cannot migrate that well from vagina to the penis From mother to child (during pregnancy, delivery and during breast feeding), the probability is 15-30% without antiretroviral therapy and with at about 1% Symptoms: Asymptomatic infection, followed by Acute phase 2-4 weeks after infection (influenza-like symptoms, aseptic meningitis) Dementia related to AIDS Opportunistic infections Retroviridae Human immunodeficiency virus (HI AIDS-related complex (ARC): prodromal (early) phase of infection with HIV that includes low fever, unexplained weight loss, diarrhea, opportunistic infections and generalized lymphadenopathy (swollen lymph glands) o Laboratory criteria separating ARC from AIDS include elevated or hyperactive B-cell humoral immune response, compared to depressed/normal Laboratory diagnostics: antibody reactivity in AIDS; follicular or mixed hyperplasia in ARC lymph nodes, leading to lymphocyte degeneration; Kaposi’s sarcoma § SINGALS THE TRANSITION TO THE FULL-BLOWN AIDS Detection of antibodies o Kaposi’s sarcoma: cancer that forms masses in skin, lymph nodes or other organs (skin lesions Detection of virus RNA are purple and flat/raised) § HHV-8 is found in the lesions of all those who are affected § Monitoring of CD4+ T-cells count Risk factors include poor immune function, either as a result of disease or specific medications, and chromic lymphedema Opportunistic infections during developed AIDS: Pneumocystis jiroveci (pneumonia) Toxoplasma gondii (encephalitis) Candidiasis Chronic herpes simplex virus (1+2) infection Retroviridae Cytomegalovirus (CMV) infection Cryptococcus neoformans infection (yeast; can cause pneumonia) Human immunodeficiency virus (HIV Mycobacterium tuberculosis Mycobacterium avium, Mycobacterium intracellulare Laboratory diagnostics: HIV encephalitis, HIV nephropathy or cardiomyopathy (=damage to these organs) Laboratory diagnostics: Detection of antibodies (direct type) à ELISA o 1st: ELISA, and if positive, 2nd: conformation via western blot www.biomedic-plze Detection of virus RNA (direct type)) à RT-PCR Monitoring CD4+ and T-cell count Antiretroviral therapy: Combination of three or more different antiretroviral drugs Usually tow nucleoside inhibitors of reverse transcriptase (NRTI) and protease inhibitor Two NRTI and one non-nucleoside inhibitor of RT, but other combinations are possible Hepatitis: Hepatitis A, B, C, D, E, G (different viruses with similar basic hepatitis symptoms) It leads to Infection and damaging of liver: icteric symptoms of jaundice and the release of liver enzymes www.biomedic-plzen. A) Hepatitis A virus Member of the PICRONAVIRUSES (naked capsid, positive sense, single-stranded RNA) Spreads by fecal-oral route with incubation period of 1 month It does not cause chronic diseases and clinically symptoms will disappear within 8 weeks Diagnostics: Serology: detection of antibodies (non-direct) RT-PCR (direct) Therapy/prevention: no antiviral therapy, vaccination possible 7  Resistant to low pH, freezing an Incubation period ca. 3 months Microbiology oral topics B) Hepatitis B virus Chronic hepatitis in 5 – 10 % of patients Member of HEPADNAVIRUSES (enveloped virus, partially very small circular dsDNA) Virion (called Dane) includes proteinkinase and polymerase enzymes It is resistant to low pH, freezing and heating Causally associated with primary hepatoc Hepatitis B antigen (HBsAg) is not a complete virion Epidemiology: Spreads parentally by needles, sexual contact or perinatally (outside the intestine) www.biomedic Incubation period approx. 3 months Chronic hepatitis occurs in 5-10% of patients Causally associated with primary hepatocellular carcinoma Diagnostics: Direct detection of HBsAg and HBeAg Detection of antibodies HBcIgM (chronic phase infection) and HBcIgG (long-term Ig) PCR of viral DNA Hepatitis www.biomedic-plzen.cz Therapy: causal therapy is available, vaccination is possible Hepatitis B virus C) Hepatitis C virus Diagnostics: Member of HEPACIVIRUS (FLAVIVIRIUSES) (enveloped, +RNA with RNA dependent RNA polymerase) It has at least 6 genotypes and more than 50 subtypes Disease: Incubating period: 4-12 weeks Direct detection of HBsAg, HBeAg Hepatitis In 80% occurs chronic disease Hepatitis Detection of antibodies D virus HBcIgM, HBcIgG Cirrhosis occurs in 5-20% of cases Development of hepatocellular carcinoma is possible Deltavirus PCR of viral DNA ssRNA Epidemiology: 71 million people are infected, 1.75 million per year and 400,000 deaths per year Laboratory diagnostics: Dependent on replication of HBsAg Detection of antibodies Causal therapy is available Present only in patients infected by h Detection of RNA by RT-PCR Vaccination (all health-care workers) Therapy: Ribavirin in combination with interferon α or other antiviral drugs Epidemiology: D) Hepatitis D virus In 4 % of patients with hepatitis B infectio Member of DELTAVIRUSES (ssRNA, dependent on replication of HBsAg which is only present Usually spread in patients infected byB virus) by hepatitis sexual contact, bloo Epidemiology: fetus In 4% of patients with hepatitis B infection, an extra infection with hepatitis D occurs Usually spreads by sexual contact, blood and from mother to fetus www.biomedic-plzen.cz E) Hepatitis E virus Member of HEPEVIRUSES (RNA, enveloped) Epidemiology: Spread by fecal-oral route Mostly common in Africa, Asia, South America Animal reservoirs are (pigs, rats, birds) www.biomedic-plzen.cz 3 million infections each year and 44,000 deaths per year Disease: Incubation period of 3-8 weeks Typical hepatitis symptoms occur Danger for pregnant women (20% mortality) 8 Microbiology oral topics 3. Urinary tract infections (UTI) and sexually transmitted diseases (STD) URINARY TRACT INFECTION (UTI): It is an infection of any part of urinary system (urethra, bladder, ureters, kidney) Classification: o Acute vs. chronic o Symptomatic vs. asymptomatic o Lower (urethritis, cystitis, prostatitis) vs. upper (pyelitis, pyelonephritis) § Inflammation of urethra à urethritis § Urinary bladder à cystitis (most frequent) Frequent urination, dysuria, strangury (=stranguria), pain in hypogastrium § Prostate à prostatitis § Kidney pelvis à pyelitis § Kidney à pyelonephritis (may be acquired from cystitis or by hematogenic way) As in cystitis + pain in lumbar region and febrile o Uncomplicated vs. complicated (in case of concrements, diabetes mellitus, immunodeficiency) Urine changes: turbidity, bad odor, hematuria (macroscopical hematuria is rare) UTI categories: Bacterial – most frequent Mycotic – less frequent (Candida) Viral – rare (Adenoviruses) Parasitic – imported only (Schistosoma haematobium) Epidemiology (individuals with predisposition to UTI): Neonates (same frequency in both sexes) Prepubertal (mainly girls) Young women Older men (>40 years; prostate hyperplasia) and women o In category 20-50, about 50x more frequent in women (shorter urethra and closer distance of rectal and urethral opening in females) People with abnormalities of urinary tract Immunocompromised patients Pathogenesis: Most common causative agents are bacteria of intestinal normal microflora (they access to urinary tract outlet via perineum) Ascendant way of infection: colonization of urethral opening à urethra à bladder à kidney o Urethra length: 4 cm in females vs. 25 cm in males Risk factors: A) Physiological and anatomical Vesicourethral reflux – urinary retention (give more time for bacterial growth in urethra) Pregnancy – compression of ureters by the uterus and progesterone-mediated smooth muscle relaxation of the bladder (incomplete emptying) Prostate hyperplasia – increased post-viding volume (amount of urine retained in the bladder after a voluntary void) Voiding dysfunctions – increased post-voiding volume B) Behavioral Sexual activity – introduction of bacteria to urinary tract (in young women causes 75-90% cystitis) C) Iatrogenic (caused by the doctor) Urinary catheterization ATB therapy – vaginal dysmicrobia, colonization by UT pathogens D) Genetic Susceptible uroepithelial cells – lower secretion of IgA (on mucous surfaces by mucous secretions) Familiar tendency 9  Symptoms of UTIs Lower UTI Microbiology oral topics Disuria = difficult urination, increase of pain in late faze of voiding (terminal disuria) Stranguria = pain (burning) in urination Symptoms of UTIs: Symptoms of UTIs Frequent urination, urge to urinate Pain in hypogastrium, suprapubic pain A) Lower Lower UTI UTI B) Upper Upper UTI UTI Disuria = difficult urination, increase of pain in late faze of voiding (terminal disuria) Symptoms of lower UTI + Stranguria = pain (burning) in urination Abdominal pain, lower back pain Frequent urination, urge to urinate Fever Nausea, vomiting Pain in hypogastrium, suprapubic pain Upper UTI Infants may have fever only + may feed less or vomit Symptoms of lower UTI + Asymptomatic bacteriuria: Abdominal pain, lower back pain FeverPositive cultivation – concentration of bacteria above the limit, no leukocyturia, biochemical parameters in normal values, no clinical sings of infection Nausea, vomiting May occur in older individuals Infants Usually, may haveno therapy fever only +with may exception feed less orofvomit pregnant women (infection of birth canal possible) and risk categories (immunodeficiency) Non-bacterial causes of UTI: Viruses – Adenovirus (blood in urine due to hemorrhagic cystitis) Fungi – Candida sp. (in elderly more frequent as pyelonephritis) Parasites – Schistosoma haematobium (helminth; parasite of blood capillaries of bladder) o only imported (e.g., Africa) o microscopic diagnosis from urine (eggs) or ELISA Causative (bacterial) agents: Most common causative agents of URI: Escherichia coli (70-90% in community), Staphylococcus epidermidis and Enterococcus spp. (10-20% in hospital each) Gram-negative rods of normal intestinal microflora: Nosocomial UTIs: Escherichia coli (most common; 80%) Nosocomial = hospital-acquired infection Proteus sp. E. coli, Klebsiella sp., Pseudomonas sp., Enterococcus sp. Klebsiella sp. Staphylococcus epidermidis (in patients with urinary catheters) Enterobacter sp. Candida sp. (fungi; cases pyelonephritis) Staphylococcus sp. Mycobacterium sp. (rare; specific diagnostic method for tuberculosis) Coagulase-negative Staphylococcus saprophyticus Diagnostics - cult Cultivation is most confident method of UTI diagnosis. There Diagnostic methods: urine specimens collected by voiding. These are bacteria wh flushed into collected urine sample. For that, test are done q A) Cultivation concentration that given limit (normal content) is evaluated Gold standard – most sensitive, most confident Culture plates are seeded by a loop calibrated for certain vol B) Microscopy surface. After culture period (24h) number of apparent colon Rapid method, low sensitivity, group identification only possible, gives information about human cell presenceconcentration per ml. (leukocytes, erythrocytes) C) Biochemical parameters of urine Supportive value only (proteins, leukocytes, erythrocytes) Diagnostics - cultivation Cultivation is most confident method of UTI diagnosis. There are always some bacteria present in urine specimens collected by voiding. These are bacteria which colonize urethral opening and are flushed into collected urine sample. For that, test are done quantitatively and recording of highe Diagnostics – specimen: concentration that given limit (normal content) is evaluated as proven UTI. Urine (specimen): Culture plates are seeded by a loop calibrated for certain volume, specimen is spread on the surface. After culture period (24h) number of apparent colonies is counted and recalculated for Changes: turbidity (high content of leukocytes), bad smell (only in bacteria with urease production -> ammonium release), concentration per ml. hematuria (macroscopically rase, microscopically common), but also without sensory detectable changes Urine specimen collection: If culture method will be employed, special attention of prevention of the urine specimen contamination must be given: o Region of urethral opening should be washed with soap and water, towel drying should not be used o Females should spread labia, males should retract preputium (foreskin) o Middle stream morning urine should be collected into sterile container (edge and cap of container should not be contaminated by fingers) o Infants: plastic pouches with adherent edges are used for collection or urine is collected from bladder by suprapubic puncture o Urine specimens should be refrigerated during storage and (fast) transport 10  Microbiology oral topics Diagnostics – cultivation: There are always some bacteria present in all urine specimens collected by voiding o Are bacteria which colonize urethral opening and are flushed into collected urine sample o For that, tests are done quantitatively and recording of higher concentration than the given limit (normal content) is categorized as proven UTI Culture plates are seeded by a loop caliper for certain volumes and the specimen is spread on the surface o After cultivation period of 24h, the number of apparent colonies is counted and recalculated for concentration per ml Colony forming unit (CFU) – estimation of the number of viable bacterial/fungal cells in a sample (CFU/mL) o One cell gives one colony in the majority of cases (but some cells do not -> dead cells or cells of bacteria which do not grow on medium) o Some cells may also adhere together and then appear as one colony § This is why the recorded concentration as CFU/mL may be slightly different than real concentration of bacterial cells o Limits for normal content = 105 CFU/mL (> means bacterial infection) § Different limits are for infants, patients with urinary catheter and patients with ATB therapy § Limit for specimens collected by puncture from bladder in healthy persons = 0 CFU/mL Diagnostic procedure: Primoculture: quantitative assay à if positive, isolation on new culture medium à identification (biochemical tests or mass spectrometry) OR Use of chromogenic culture media (both quantitative assay and identification in primoculture) o Chromogenic media contain a set of species + specific substrates, each labeled by different chromogen o Utilized substrates stain colonies of bacteria by specific color o Spectrum of used substrates covers spectrum of pathogenic species for given purpose (here UTI) o Diagnostic cycle is faster and acceptably confident Other methods: A) Rapid tests Diagnosis Dip-stick methodsother from freshlymethods voided urine: detection of hematuria, leukocyturia, leucite-esterase apid tests o Dip-stick tests may indicate possible UTI, but have low sensitivity p-stick methods, from freshly voided urine: detection of hematuria, lecocyteuria, leucocyte-esterase p-stick tests may indicate possible UTI, but has low Positive results also in non-UTI diseases (cancers, intoxication, kidney dysfunctions) § sensitivity ositive results also in non-UTI diseases (cancers, intoxication, kidney dysfunctions icroscopy B) Microscopy etection of leucocytes, erythrocytes, bacteria Detection of leukocytes, erythrocytes, bacteria dicates possible UTI, but has lower sensitivity than cultivation, causative aget identification is not possible Indicates possible UTI, but has lower sensitivity than cultivation (causative agent identification is not possible) Urine with leucocytes and G-neg. rods, Gram stain C) Urinary tract imaging rinary tract imaging Used as an additional method to cultivation in complicated UTIs (concrements, obstructions or cancers) an additional method to cultivation in complicated UTI (concrements, obstructions, cancers) CAUSATIVE AGENTS Staphylococcus sp.: Coagulase negative ones Diagnosis: Microscopy: Gram-positive cocci with formation of clusters Cultivation: white colonies (no pigment production) and in blood agar NO hemolysis Tests: catalase positive and plasmacoagulase negative (coagulase negative Staphylococcus) Exact species identification is possible by other biochemical tests or by mass spectrometry Enterobacteria of normal intestinal microflora: Gram-negative rods Diagnosis: Microscopy; species identification by biochemical tests or by mass spectrometry is possible Cultivation: o Blood agar (light gray color with no pigments) o Endo agar (lactose positive) o NO hemolysis (or only weak non-alpha, non-beta o XLD (fermentation of saccharides positive) 11  Microbiology oral topics Proteus sp.: Gram-negative rods Member of normal GIT microflora Diagnosis: Microscopy; species identification also by biochemical tests or by mass spectrometry Cultivation: o Blood agar (full motility; they show crawly growth) o Endo agar (motility partially inhibited, lactose negative) o XLD (motility fully inhibited, weak fermentation of saccharides) STD most common caus Candida sp.: Fungi with oval cells, usually blue (dead cells are red) Diagnosis: Bacterial Viral Microscopy; species identification also by biochemical tests or by mass spectrometry Treponema pallidum (syphilis) HIV (huma Cultivation: Sabouraud agar (large white matt colonies) à it is typically used for fungi Diagnostic - cultivation SEXUALLY TRANSMITTED DISEASES (STD): Neisseria gonorrhoeae (gonorrhea) Chlamydia trachomatis Herpes sim HPV (Hum Mycoplasma, Ureoplasma Hepatitis B nostic procedure: Are infections that are commonly spread by sexual activity (vaginal, oral or anal intercourse) STD most STDcommon most common causative causative agentsagents MCV (moll oculture: quantitative assay Besides sexual activity, some STD may also spread by: i.v. drug abuse, blood transfusion or congenital way (present from birth) Most common causative agents: Bacterial Bacterial Viral Viral Parasitic Treponema pallidum (syphilis) atively useTreponema of chromogenic culture media pallidum (syphilis) HIV (human immunodeficiency virus) HIV (human immunodeficiency virus) Trichomonas vaginalis Neisseria gonorrhoeae (gonorrhea) Herpes simplex virus 1,2 (herpes genitalis) Neisseria gonorrhoeae (gonorrhea) Herpes simplex virus 1,2 (herpes genitalis) quantitativeChlamydia assay trachomatis and identification in primoculture. Chlamydia trachomatis Chromogenic HPV (Human Papillomavirus) media contain set of species Phthirus pubis Mycoplasma, Ureoplasma HPV (Human Papillomavirus) Hepatitis B virus fic substrates, each labeled by Mycoplasma, Ureoplasma different chromogen. Utilized substrates stain Hepatitis B virusMCV (molluscum contagiosum virus) colonies of bacteria ecific color. Spectrum of used substrates covers spectrum MCV (molluscum contagiosumof pathogenic virus) species for given purpose UTI). Diagnostic cycle isParasitic faster and acceptably confident. CAUSATIVE AGENTS: Parasitic Trichomonas vaginalis Phthirus pubis Trichomonas vaginalis Phthirus pubis Neisseria gonorrhea (gonorrhea): Gram-negative diplococcus Incubation: 1-15 days (usually 2-5 days) Males (urethritis): stanguria (painful, frequent urination of small volumes), urethral discharge, rarely asymptomatic Females (urogenital infection): infection of urethra and endocervical canal; dysuria (painful/difficult urination), vaginal discharge and bleeding (between periods), pelvic pain, often asymptomatic (70%) In both sexes also oral and rectal primary infection possible Conjunctivitis in newborn (transmission via delivery) or in children (sexual abuse) Disseminated gonorrhea: Affects about 1% of affected Hematogenous spread to the skin, tendons, joints Symptoms: fever, chills, arthritis (especially of large joints), hemorrhagic pustules on the acral parts (e.g., toes or fingers) Neonatal gonorrhea: Infection of newborn during delivery in the infected birth canal Leads to ocular infection – conjunctivitis: purulent discharge from conjunctiva, swelling of eyelids o If left untreated: damage of cornea – blindness, but also arthritis, meningitis, or sepsis Therapy should be administered to both mother and newborn Prophylaxis: solution of antisepticum is instilled into conjunctiva to all newborns 12 Microbiology oral topics Diagnosis: A) Cultivation Diagnosis of gonorrhea Specimens collection by urethral swab (basic, rectal, throat swabs, swab from conjunctiva, synovial fluid in disseminated gonorrhea) Microscopy Fresh samples and short-term storage at room temperature Gram stained smears of urethral swabs Medium is a selective modification of chocolate agar (antibiotics and antimycotics are Rapid, low added for inhibition of conducting normal microflora) sensitivity Confirmation by biochemical tests B) PCR Sensitive and no need of fresh specimens It is the standard diagnostic method C) Microscopy Gram-stained smears of urethral swabs Rapid, but low sensitivity Gram-negative diplococci in cytoplasma of leucocytes (hit-micrscopewb.hc.msu.edu) Treponema pallidum (syphilis): Gram-negative, helical rods, motile, group spirochaete They are found only extracellular Causes syphilis, other subspecies cause non-veneric infections (no STD) Virulent strains have slimes (other layer of soluble polysaccharides) Syphilis – clinical stages: A) Primary Occurs 2-6 weeks after infection Syphilis clinical stages secondary Formation of ulcer (chancre = ulcus durum) in place of entry (genital, rectal, oral or other side) 4 weeks 2 years after infection Ulcer: firm painless, non-itching, clean base, sharp borders; regional lymphadenitis possible Various manifestation: skin, mucous membranes, lymf nodes B) Secondary Comonly: non-itchy rash on the trunk and extremities, palmes, depigmentation Other symptoms: fever, sore throat, malaise, weight loss, and headache Occurs 4 weeks to 2 years after infection Maculopapular or pustular rash Syphilis clinical stages Tertiary o Various manifestations: skin, mucous membranes, lymph nodes May develop 3-15 years after infection, latent phase between secondry and tertiary stage Commonly: non-itchy rash on the trunk, extremities and palms Three + depigmentation forms: Other symptoms: fever, sore throat, malaise, weight loss and headache C) Tertiary Syphilis clinical stages Gummatous syphilis (15%) tumor-like balls on skin with necroses (also bones or other tissues) Neurosyphilis (7%) involvement of CNS syphilitic meningitis, paresis Tertiary Cardiovascular syphilis (10%) syphilitic aortatis, aortal aneurysma May develop 3-15 years after infection, latent phase between secondry and tertiary stage clinicalgate.com Three forms: Gummatous syphilis (15%) tumor-like balls on skin with necroses (also bones or other tissues) Neurosyphilis (7%) involvement of CNS syphilitic meningitis, paresis Cardiovascular syphilis (10%) syphilitic aortatis, aortal aneurysma Congenital syphilis Conge Transplacenthal infection or during birth Transplacenthal infection or during birth Congenital syphilis: At any stage of infection in mother At any stage of infection in mother Transplacental infection or during birth which can occur at any stage of infection May cause in theormother stilborn or abortion range of serious manifestations in infants: anemia, May cause stilborn or abortion or range hepatosplenomegaly, pulmonarysyphilis, Gummatous inflamation, bones and joints deformations, dental defects www.sciencephoto.com hepatosplenomegaly, pulmonary inflama among others May cause stillborn, abortion or ranges of serious manifestations in infants: anemia, hepatosplenomegaly, pulmonary inflammation, bones andothers among joints Seroscreening of pregnant women ATB therapy may prevent congenital infection deformation, dental defects Seroscreening of pregnant women ATB Seroscreening of pregnant women is possible + ATB therapy may prevent congenital infection Diagnosis: Gummatous syphilis, www.sciencephoto.com A) Indirect immunology (antibody detection = serology) Standard diagnostic method Early evidence - bullae and vesicular rash Late evidence - notched incisors -Hutchinson teeth Non-culturable bacteria (antigen of T. pallidum is limited) (www.sciencephoto.com) Non-treponemal test (beef cardiolipin is used as an antigen for the first test; test has low specificity but sufficient sensitivity) Early evidence - bullae and vesicular rash Treponema test (antigen of T. pallidum is used for confirmation after positive non-treponemal test; usually by Western blot for IgG and IgM) B) Direct detection Possible in primary stage only Specimen: swab from the ulcer Methods: Microscopy (native mounts; dark field microscopy), PCR, direct immunofluorescence 13  Microbiology oral topics Chlamydia trachomatis: Strictly intracellular, culturable in tissue cultures only A) Serotype = Serovars D-K Urogenital infections – urethritis, cervicitis, pelvic inflammation disease, prostatitis, epididymitis Often asymptomatic (50-80%) May result in infertility in both males and females B) Serovars L1-L3 Lymphogranuloma venereum – spread in lymphatic nodes, swelling and rupture Occurs in tropical regions and since 2003 increased evidence in Europe C) Serovars A-C Non-veneric Trachoma – infection of the eyes, causing blindness Occurs in tropical regions with no occurrence in Europe Diagnosis: Strictly intracellular bacteria which are non-culturable on artificial medias Specimens: urethral swabs, cervical swabs, first-voided urine A) PCR B) Cultivation on tissue cultures Conventional PCR is the currently used method Less sensitive, but 100% specific Real-time PCR (qPCR) is the most confident quantitative method Expense and slow method Used in special indications C) Serology Detection of genus or species-specific antibody Available as complement fixation test, ELISA or immunofluorescence (are widely used methods, but have limited confidence) Trichomonas vaginalis (trichomoniasis): Protozoan parasite (protist of flagellate group) Non-invasive parasite of mucosal surfaces and possible erosion of epithelium In women: vaginal infection and itching, pain during sex, malodorous yellow-green discharge In men: asymptomatic, rarely mild urethritis Infections left undetected/untreated may persist for years and have the tendency of symptoms moderation in time with staying infective (all sexual partners should be treated) Diagnosis: A) Microscopy Limited sensitivity Specimen: vaginal swab or urethral swab in men (males are asymptomatic, so test is rarely done) Dry smears, methanol fixed, and Giemsa stain is used Usually, a second slide by Gram stain is used (symptoms of trichomoniasis are not specific and microscopy test may detect also material and mycotic Vaginal smears - microscopy Vaginal smears - microscopy Vaginal smears - microscopy infections, including non-veneric (no STD) ones, which are better detectable on Gram-stained slides) Mycotic infection yeast, G+ rods, leucocytes, epithelial cells, Gram stain Gonorrhea Leucocytes, Neisseria gonorrhoeae, Gram stain Bacterial infection, nonpurulent epithelial cells, various bacteria (leucocytes not present) Note: gonorrhea is rarely detected in vaginal swabs, if suspected, urethral swabs should be collected () (www.dowdiagnostics.com) B) Cultivation Optional method with higher sensitivity which may be the only possibility for diagnosis in chronic patients with low content of parasite cells in smears (not detectable by microscopy) Use of liquid culture medium C) PCR Most frequent method 14  Microbiology oral topics Questions: 1) Chocolate agar contains: Blood; mixing temp. of agar and blood is 80°C (erythrocytes are destroyed) o Some bacteria need the contents of erythrocytes (e.g., Neisseria gonorrhea, Streptococcus pneumonia, Hemophilus influenzae) 2) Mycobacteria in Gram-stain appear: Red 3) Blood agar is: Both a selective and differential (diagnostic) medium 4) Alpha hemolysis is: Incomplete hemolysis 5) Direct immunological methods detect: Antigen 6) Microaerophilic bacteria: Grow at low concentration of oxygen and do not tolerate its absence 7) Gram-positive bacteria appear in Gram stain: Blue 8) Candida is: Fungus 9) If procedure of urine specimen collection or its storage is wrong, there is a risk of: False positive result 10) What is the correct temperature condition for urine specimen storage for the purpose of UTI diagnosis? Refrigeration 11) What causes UTI? E. coli, Staphylococcus saprophyticus, Staphylococcus epidermidis, Enterococcus sp., (G+), Pseudomonas aeruginosa (G-) 4. Gastrointestinal tract infections Classification: GIT infections common causative agents Stomach à gastritis Bacterial Viral Small intestine à enteritis (most frequent) Salmonella enterica Rotaviruses Large intestine à colitis Campylobacter jejuni Adenoviruses The latter is often associated with enterocolitis or gastroenteritis Yersinia enterocolitica Noroviruses Shigella Primary source of infection: zoonoses from tissues of infected sp.(undercooked meat) or aminal products animals Astroviruses (egg, milk) Helicobacter pylori Secondary source of infection: food contamination during proceeding (fecal-oral transmission) = antroponose (human to human transmission) Escherichia coli infections common GIT infections causative common causative agents Alimentary intoxication (= food poisoning) is not an infection, but the intoxication is caused by microbial toxins produced and accumulated in contaminated food agents Common causative agents: acterial Bacterial Viral Viral Parasitic Salmonella enterica Rotaviruses Enterobius vermicularis almonella enterica Rotaviruses Campylobacter jejuni Adenoviruses Giardia intestinalis ampylobacter jejuni Yersinia enterocolitica Adenoviruses Noroviruses ersinia enterocolitica Shigella sp. Noroviruses Astroviruses Helicobacter pylori higella sp. Astroviruses Escherichia coli elicobacter pylori scherichia coli Diagnostic Parasitic methods: Enterobius vermicularis Viral: antigen detection (direct immunological methods), PCR Giardia intestinalis arasitic Bacterial: cultivation (microscopy for identification), PCR nterobius vermicularis Parasitic: microscopy, antigen detection, PCR iardia intestinalis 15 Microbiology oral topics CAUSATIVE AGENTS: Salmonella enterica: Gram-negative rod, group of Enterobacteriaceae Taxonomy: Among 6 subspecies of Salmonella enterica, only Salmonella enterica, subspecies enterica may cause human infection There are many serotypes of this subspecies, e.g., Salmonella enterica subsp. enterica Typhimurium Diseases: Salmonellosis (= non-typhoidal salmonellosis) caused by zoonotic serotypes; prevalent in Europe Typhoid fever – caused by anthroponotic serotypes Typhi and Paratyphi (in Europe important only) Salmonellosis (non-typhoidal): Diagnostic of salm Cultivation Signs and symptoms: Incubation of 10-36h (typically 10-12h); fever, vomiting, abdominal cramps, profuse diarrhea (watery, non-bloody, typically lasts 4-7 days Specimen: stool or rectal swab Complications: Selective and dipherential culture media inhibition of part dipherentiation of pathogenic bacteria (MacConkey, Endo, TSI o Serious dehydration – may require hospitalization, i.v. fluid deliver Identification of serotype by agglutination with specific anti o Extraintestinal spread – osteomyelitis, meningitis, endocarditis, soft tissues abscess, sepsis o Infants and elderly are at risk Diagnostic of salmonellosis Cultivation Cultivation: Specimen: stool or rectal swab Selective and dipherential culture media inhibition of part of normal microflora (G+ bacteria), Specimen: stool or rectal swab dipherentiation of pathogenic bacteria (MacConkey, Endo, TSI, XLD) Identification Selective and differential culture media (inhibition of part of normal of serotype microflora by agglutination à Gram-positive with specific antibodies bacteria) S. enterica on Endo agar and XLD agar Serotype Differentiation of pathogenic bacteria (MacConkey, Endo, TSI, XLD) Identification of serotype by agglutination with specific antibodies Widal test Detection of Ab in serum by agglutination with dead bacteria Widal test: detection of antibody in serum by agglutination with dead bacteria and their cellular fractions Salmonella enterica serotypes Typhi and Paratyphi: They cause typhoid fever (in Europe rare) S. enterica on Endo agar and XLD agar Serotype identification by agglutination Antroponose – human-to-human transmission, no animals involved Widal test They enter the lymphatic system, leading to a systemic formDetection of the disease of Ab in serum by agglutination with dead bacteria and their cellular fractions The endotoxins influence the vascular and nervous system, leading to fluid disbalance, sepsis or septic shock Symptoms: vomiting, febrile, headache, diarrhea or constipation Vaccine is recommended prevention for travelers and is controlled in regions of occurrence Shigella dysenteriae: Gram-negative rod, group Enterobacteriaceae Genetically related to Escherichia coli Types: S. dysenteriae, S. sonnei, S. flexneri, S. boydii Shiga toxin: Interacts with ribosomes à inhibition of proteosynthesis à cell death Damage of enterocytes à invasion to bloodstream Damage of endothelium in blood vessels à damage of endothelium in glomeruli à proteinuria, hematuria à hemolytic uremic syndrome Enterohemorrhagic strains of E. coli have similar toxins (shiga-like toxin) Transmission and symptoms: Fecal-oral route (not from animals) à causes SHIGELLOSIS Mild abdominal discomfort to serious dysentery with profuse diarrhea, fever, vomiting, abdominal cramps o Dysentery = intestinal inflammation, primarily in colon; type of gastroenteritis that results in diarrhea with blood Complications: Diagnostics of shigellosis Serious dehydration, mucosal ulceration, rectal bleeding, hemolytic uremic syndrome Cultivation identical method as in salmonellosis ATB therapy may be problematic in multiresistant strains Differentiation: Salmonella positive for H2S production, Shigella H2S negative It is globally one of the most frequent bacterial cause of GIT infection (inSpecies Europeidentification significantly less frequent by biochemical teststhan salmonellosis) or mass spectrometry Cultivation: Identical method as in salmonellosis Differentiation: Salmonella positive for H2S production, Shigella H2S negative Species identification by biochemical tests or mass spectrometry Shigella sp. on Endo agar and XLD agar 16 Microbiology oral topics Yersinia enterocolitica: Gram-negative rod, group Enterobacteriaceae Zoonose – pigs, pork meat or contaminated water Incubation of 3-7 days Symptoms: Watery diarrhea which may be with blood, dissemination into mesenteric lymph nodes, right lower abdominal pain (may mimic appendicitis) Range of symptom intensity, from mild to similar as with salmonellosis or shigellosis Diagnosis: Stool sample, cultivation on selective-differential media, identification by biochemical test or mass spectrometry, hemocultures in complicated forms Escherichia coli: Gram-negative rod, group Enterobacteriaceae Part of GIT normal microflora of mammals Most normal flora E. coli are non-pathogenic in GIT Strains of E. coli may be equipped by virulence factors enabling to cause infections (virulent strains) Virulent strains may differ significantly in characteristics of the diseases (incubation period, pathogenesis, symptoms, epidemiology) Virulent strains: Enteropathogenic E. coli (ETEC) – occurs in developing countries and is a traveler’s diarrhea Enterohemorrhagic E. coli (EHEC) – is a shiga toxin-producing E. coli (STEC); shiga-like toxin, can cause dysentery, hemolytic uremic syndrome and kidney failure Enteropathogenic E. coli (EPEC) – causes neonatal and infant diarrhea, also as HAI (hospital acquired infection) in neonatal and pediatric units Enteroinvasive E. coli (EIEC) – causes dysentery, globally prevalent strain Transmission: Majority by fecal-oral route, some strains also zoonotic with various reservoirs (wild, farm and pet animals) Diagnosis: Isolation of E. coli by cultivation; virulent strains cannot be identified by any biochemical test nor by mass spectrometry o NOT detectable by XLD Strain identification by antigenic properties – agglutination with specific antibodies PCR is available for some strains Vibrio cholerae: Gram-negative, curved rod, flagellated (motile) Cause cholera disease – acquired from contaminated water, seafood (contaminated marine water) and fecal-oral route Cause gastroenteritis (inflammation of stomach and intestines) or choleratoxin – release of Cl- ions and water into intestinal lumen o Causes profuse diarrhea, liquid loss together with vomits up to 40L per day, release of diarrheal stool may not be under control of the patient Symptom intensity is dependent on V. cholerae strain (phenotype) – some types cause mild infections or may be asymptomatic Diagnosis: cultivation of selective media It is a globally important pathogen; occurrence in local epidemies in crowded places with limited hygiene and/or contaminated water supply (refugee camps) Vaccination: recommended for travelling into regions with cholera occurrence Campylobacter: Gram-negative, curved rod, flagellated (motile), microaerophilic It is adapted to GIT of warm-blooded animals Campylobacter jejuni – most common, found in GIT of birds, poultry is main source of infection (usually by chicken meat) Other species: C. coli (pigs), C. fetus (cattle) Disease: Transmission: In animals usually asymptomatic Foodborne: insufficiently cooked meat, unpasteurized milk (main way) In humans: enteritis (inflammation of small intestine) or enterocolitis Fecal-oral (human-to-human) (inflammation of intestine and colon) Waterborne (contaminated water) 17 Microbiology oral topics Campylobacteriosis: Incubation period of 2-11 days In Europe increased incidence in the last two decades Symptoms: Prodromal phase – febrile, headache, myalgia After 12-24 hours – abdominal pain, cramps, diarrhea, dysentery (gastroenteritis with bloody diarrhea) The symptomatic phase lasts typically Helicobacter pylori Usually, no vomiting Gram-negative, curved rod, flagelated - motile, microaerofilic Relative to Campylobacter (former name Campylobacter pylori) Complications: toxic megacolon, invasion of bloodstream (risk of sepsis) Adaptation for survival in stomach: local inhibition of HCl production, urease enzyme production Diagnosis: of amonium neutralisation of pH Cultivation of a stool sample, detection requires special selective Specific pathogen culture of human, medium, 30-60%, prevalence cultivation must beasymptomatic majority done under microaerophilic atmosphere PCR Linked to: Chronic gastritis Helicobacter pylori: Gastric or duodenal ulcers Gastric carcinoma Gram-negative, curved rod