Clinical Bacteriology - Spirochetes & Miscellaneous Bacteria PDF

BS MLS 3A DEANNASAURSAAAAAAA When reproducing, a spirochaete will undergo asexual transverse binary fission. In addition, the spirochete...

BS MLS 3A DEANNASAURSAAAAAAA When reproducing, a spirochaete will undergo asexual transverse binary fission. In addition, the spirochetes are microaerophilic or anaerobic and are extremely sensitive to The spirochetes are all long, slender, helically curved, gram- oxygen toxicity. The complete genome sequence has revealed there negative bacilli, with the unusual morphologic features of axial fibrils are no genes for catalase or superoxide dismutase. The order of and an outer sheath. Spirochaetales is divided into two families: Treponema appear as slender with tight coils; Borrelia are somewhat thicker with fewer and looser coils; and Leptospira 1. Spirochaetaceae resembles Borrelia except for their hooked ends 2. Leptospiraceae Chlamydia and Rickettsia are obligate intracellular parasite Two of the four genera of Spirochaetaceae, Treponema and Borrelia, include species that are pathogenic to man. Among Leptospiraceae, only one genus, Leptospira, has pathogenic species. Spirochaetes (also spelled spirochetes) belong to a phylum of distinctive diderm (doublemembrane) bacteria, most of which have long, helically-coiled (corkscrew-shaped) cells. Spirochaetes are chemoheterotrophic in nature, with lengths between 5 and 250 μm and diameters around 0.1–0.6 μm. They are distinguished from other bacterial phyla by the location of their flagella, sometimes called axial filaments, which run lengthwise between the bacterial inner membrane and outer membrane in periplasmic space. These cause a twisting motion which allows the spirochaetes to move about.  BS MLS 3A DEANNASAURSAAAAAAA The causal agent of Syphilis, Treponema pallidum, was first discovered in 1905 by a German scientist Schaudinn in the primary sores (Chancres, pronounced shankers) of a syphilitic patient. In electron micrograph, T. pallidum is seen to be covered by an outer periplast which covers the whole organism when the periplast is removed by digestion with trypsin or pepsin, the fine filaments are seen twisted around the organism. It seems likely that they are contractile and maintains the characteristic shape of motility of the organism. T. pallidum shows rotary corkscrew like highly characteristic motility (Angulation) which can be easily seen under dark field microscope.  Class – Spirochaetes  Order – Spirochaetales  This organism has not been successfully cultured in vitro.  Family – Spirochaetaceae  Preservation media: 18-21 days.  Genus – Treponema  Dieterle stain: Can be used to visualize T. pallidum.  Species – Treponema pallidum  T. pallidum cannot be cultivated artificially, but the pathogenic  strain (Nichol’s strain) can be grown in the testicles of experimentally inoculated rabbit.  Certain other non-pathogenic strains (e.g. Reiter strain) can be cultivated under strict anaerobic condition in Smith Noguchi  Treponemes are thin, delicate, helically-coiled, corkscrew- medium. shaped organisms.  They are microaerophilic and actively motile.  They are measures about 10 to 14 micrometers long and 0.1 to 0.2 micrometer wide. Treponema induces at least 3 types of antibodies:  They have 8-24 sharp and angular spirals, at regular intervals a) Reagin antibodies: react in standard or non-specific test for of about 1 micrometer. syphilis.  T. pallidum can be stained by Giemsa stain in 1: 10 dilution over b) Group antigen: found in pathogenic and non-pathogenic a long period, it appears pink in color in contrast to the purplish treponemes. color of non-pathogenic spirochaete. c) Polysaccharide antigen: species-specific, demonstrated by  The organism can be demonstrated by India ink or by Fontana’s specific T. pallidum tests. silver impregnation method using the exudate from the chancre.  In tissues, spirochaetes can be stained by Levaditi’s silver impregnation method in which silver salts are allowed to  It caused Syphilis disease. penetrate the inside of the cells.  Humans are the only natural host for T. pallidum and infection  Metallic silver is then precipitated inside the spirochaetes by occurs through sexual contact. means of a reducing solution. The organism appears black  The organism penetrates the mucous membrane or enters against a yellowish black background. minuscule breaks in the skin.  BS MLS 3A DEANNASAURSAAAAAAA  In the women, the initial lesion is usually on the labia, the walls 3. Tertiary Syphilis: of the vagina, or the cervix.  In the tertiary syphilis, the spirochaetes become localized  In men, it is on the shaft or glans of the penis. producing low grade inflammatory lesions in the cardiovascular  A chancre (a small hard painless nodule at the site of entry of and central nervous systems and in chronic granulomata pathogen) also may occur on lips, tongue, tonsils, (gummata) in the skin, bone and internal organs. This is also o anus, or other skin areas. called chronic stage. Thus, tabes dorsalis and meningo,  Perivascular inflammation: This typically consists of a cardiovascular syphilis may be detected 5-15 years after the proliferation of adventitial cells; perivascular cuffing with: appearance of primary chancre. o lymphocytes, monocytes, and plasma cells; and  The late symptoms of syphilis depend on the organ that is swelling and proliferation of endothelial cells. attacked by the spirochaetes. The most frequent, serious and disabling results of syphilitic infection are disease of heart, arteries, causing sac-like dilatation called aneurysm. 1. Primary Syphilis  These may burst and the patient may die of the hemorrhage.  T. pallidum gains entrance into the body through the skin and Syphilis is one of the principle causes of diseases of the aortic mucous membrane. valves, but not responsible for arteriosclerosis; and cerebral  During sexual intercourse, it generally infects genitalia, hemorrhage may occur in young person. occasionally the lip during kissing; and it is not homosexually  In untreated cases, T. pallidum lodges in the central nervous acquired. system (brain and spinal cord) where they cause a chronic  After an incubation period of 4-6 weeks, the initial sore of inflammation (Neurosyphilis) and destroy the nerve tissue. primary syphilis appears on the genitalia. Various forms of insanity result. These lesions result in paresis  This lesion is at first small red purple (chancre), it gradually (general paralysis) and locomotors ataxia or tabes. enlarges, becomes indurated and necrosis in the center of an  In paresis, spirochaetes are present in the tissue of the brain; ill ulcer; then a generalized lymphadenopathy of the inguinal gland tabes, in the spinal cord. develops. Spirochaetes are present in large numbers in the exudate of the primary chancre. 4. Congenital Syphilis:  As the sore heals, the organism becomes less numerous and  T. pallidum can cross the placental barrier and a syphilitic may not be demonstrated in the exudate, but, at this stage, it mother especially in secondary syphilitics—may transmit the may be possible to find them in fluid aspiration by lymph gland infection to her fetus. puncture.  The lesions of congenital syphilis are similar to those already described when the infection is massive, the child may be 2. Secondary Syphilis: stillborn or survive for a short time.  After 6 to 12 weeks of the appearance of primary chancre, the  The most important defects of late congenital syphilis are secondary syphilis develops with the invasion of blood stream mental deficiency, chronic meningitis, blindness and deafness. and widely distributed through-out the body.  The clinical manifestations in this stage are fever, a generalized roseolar skin rash, mucous patches in the mouth and  T. pallidum-pertenue. condylomata of the anus and vulva.  T. carateum: the non-sexually transmitted treponematoses,  The severity of the lesion is extremely variable. Sometimes the yaws and pinta. lesions heal up and disappear completely.  BS MLS 3A DEANNASAURSAAAAAAA Two species: Two species of Borrelia of importance in humans:  L. interrogans – parasitic  B. recurrentis causes relapsing fever  L. biflexa - contains free-living organism  B. burgdorferi causes Lyme disease Within the species interrogans there are several different serogroups and serovars responsible for disease in humans and  Less finely coiled than the leptospires. Cells 0.2-0.5 μm in animals. diameter; stain readily, so it is visible by light microscopy.  B. recurrentis spread from person to person by lice. Lyme disease is a zoonosis transmitted to humans by hard ticks  Finely coiled spirochetes with hooked ends. Cells 0.1-0.2 μm in (Ixodes spp.). diameter, up to 20 μm in length. Not visible by direct light microscopy unless stained by silver impregnation or immunofluorescent methods. Dark ground microscopy reveals  In relapsing fever, the relapsing element may be due to antigen rotational and directional motility by means of periplasmic switching. Lyme disease slowly progressive rather than flagella. relapsing. Characteristic skin lesion ‘erythema chronicum migrans’ occur in approximately 50% of cases. Joint pains and fatigue common and later, in untreated cases, neurologic and  Leptospirosis in humans is a zoonosis, usual host being cardiac manifestations. rodents, bats, cattle, sheep, goats and other domestic animals. Leptospires excreted in urine contaminate food and water. Infection occurs by contact with either through occupation or  B. recurrentis demonstrated in blood smears by staining with recreation. Organisms may penetrate unabraded skin and Giemsa or acridine orange. conjunctiva.  B. burgdorferi much more difficult to visualize. Culture from biopsy material possible, but difficult; diagnosis usually by serology.  Leptospirosis or Weil’s disease in humans and animals.  Direct microscopy of blood and urine possible, but difficult to interpret. Leptospira can be grown, with difficulty, in special serum-containing media. Serologic diagnosis is usual. Rickettsiae are obligate intracellular parasites. They are the agents of:  Typhus  Spotted fevers  Q fever  BS MLS 3A DEANNASAURSAAAAAAA b) Phase II – organisms are produced by repeated passage in  In the United States, there are two rickettsial diseases of culture, are nonvirulent, and have lost the ability to significance: synthesize certain surface antigens. o Rocky Mountain spotted fever caused by Rickettsia  The clinical importance of phase variation is that patients with rickettsii chronic Q fever have a much higher antibody titer to phase I o Q fever caused by Coxiella burnetii antigens than those with acute Q fever.  Several other rickettsial diseases such as epidemic, endemic, and scrub typhus are important in developing countries.  Rickettsialpox, caused by Rickettsia akari, is a rare disease  The most striking aspect of the life cycle of the rickettsiae is that found in certain densely populated cities in the United States. they are maintained in nature in certain arthropods such as ticks, lice, fleas, and mites and, with one exception, are transmitted to humans by the bite of the arthropod.  Rickettsiae are very short rods that are barely visible in the light  The rickettsiae circulate widely in the bloodstream microscope. (bacteremia), infecting primarily the endothelium of the blood  Structurally, their cell wall resembles that of gram-negative vessel walls. rods, but they stain poorly with the standard Gram stain.  The exception to arthropod transmission is C. burnetii, the  Rickettsiae are obligate intracellular parasites, because they cause of Q fever, which is transmitted by aerosol and inhaled are unable to produce sufficient energy to replicate into the lungs. extracellularly.  Virtually all rickettsial diseases are zoonoses (ie, they have an  Therefore, rickettsiae must be grown in cell culture, animal reservoir), with the prominent exception of epidemic embryonated eggs, or experimental animals. typhus, which occurs only in humans.  Rickettsiae divide by binary fission within the host cell, in  It occurs only in humans because the causative organism, R. contrast to chlamydiae, which are also obligate intracellular prowazekii, is transmitted by the human body louse. parasites but replicate by a distinctive intracellular cycle.  Several rickettsiae, such as: 1. Rickettsia prowazekii 2. Rickettsia tsutsugamushi 3. R. rickettsii They possess antigens that cross-react with antigens of the OX strains of Proteus vulgaris.  The Weil-Felix test, which detects anti-rickettsial antibodies in a patient's serum by agglutination of the Proteus organisms, is based on this cross-reaction.  C. burnetii has a spore-like stage that is highly resistant to drying, which enhances its ability to cause infection.  It also has a very low ID 50, estimated to be approximately one organism.  C. burnetii exists in two phases that differ in their antigenicity and their virulence: a) Phase I – organisms are isolated from the patient, are virulent, and synthesize certain surface antigens.  BS MLS 3A DEANNASAURSAAAAAAA  Elementary bodies are usually present in the semen of infected men and vaginal secretions of infected women.  When they come into contact with a new host cell, the elementary bodies bind to the cell via interaction between adhesins on their surface and several host receptor proteins and heparan sulfate proteoglycans.  Once attached, the bacteria inject various effector proteins into the host cell using a type three secretion system.  These effectors trigger the host cell to take up the elementary bodies and prevent the cell from triggering apoptosis.  Within 6 to 8 hours after infection, the elementary bodies transition to reticulate bodies and a number of new effectors are synthesized.  These effectors include a number of proteins that modify the inclusion membrane, called Inc proteins, as well as proteins that redirect host vesicles to the inclusion.  8 to 16 hours after infection, another set of effectors are synthesized, driving acquisition of nutrients from the host cell.  At this stage, the reticulate bodies begin to divide, causing the inclusion to expand.  If several elementary bodies have infected a single cell, their Members of the genus Chlamydia are Gram-negative bacteria. inclusions will fuse at this point to create a single large inclusion While the majority of these bacteria have a thin peptidoglycan, the in the host cell. peptidoglycan of these species is not easily detectable.  From 24 to 72 hours after infection, reticulate bodies transition to elementary bodies which are released either by lysis of the In C. pneumoniae and C. trachomatis, however, genomic host cell or extrusion of the entire inclusion into the host genital studies have shown that they encode for proteins that may be involved tract. in the synthesis of peptidoglycan. In addition, they encode for a major outer membrane protein, a protein located on the surface of C. trachomatis and C. psittaci. This is an important protein used for serological identification of different forms of C. psittaci and C. trachomatis. The absence of peptidoglycan in Chlamydia bacteria has been suggested to be an evolutional adaptation that allows them to effectively invade and thrive in the cells of their respective hosts. Infective and reproductive forms of Chlamydia include elementary bodies (EB) and reticulate bodies.  BS MLS 3A DEANNASAURSAAAAAAA o Neonatal conjunctivitis and other infections: o Over 50 percent of infants born to women infected with  Psittacosis, also known as ornithosis, denotes a zoonotic Chlamydia trachomatis, serotypes D–K (animal) disease that is transmitted to humans by inhalation of o The most common presentation is inclusion conjunctivitis of dust contaminated with respiratory secretions or feces of the newborn. infected birds. o If untreated, the infection can lead to permanent scarring of  The human disease usually targets the lower respiratory tract. the cornea or conjunctiva  There is an acute onset of fever, hacking dry cough, and flulike symptoms.  Inclusion conjunctivitis in adults:  Bilateral patchy pulmonary infiltrates are observed. o Individuals of any age may develop transient purulent  Enlargement of liver and spleen is a frequent accompanying conjunctivitis caused by trachomatis serotypes D–K. feature. o Such individuals are often found to be genitally infected as well.  Non-gonococcal urethritis: o Chlamydial NGU is symptomatically similar to infections  Chlamydia pneumoniae is a respiratory pathogen causing caused by Neisseria gonorrhoeae. pharyngitis, sometimes followed by laryngitis, bronchitis, or o Whether locally symptomatic or not, the infection may interstitial pneumonia. ascend into the upper reproductive tract to involve the  It is a significant cause of community-acquired respiratory epididymis in men and fallopian tubes and adjacent tissues infection, occurring worldwide and without seasonal incidence. in women (pelvic inflammatory disease).  Lymphogranuloma venereum: o Chlamydia trachomatis, serotypes L1, L2, and L3, cause  for the chlamydial infections ocular, urethral, vagina cervical lymphogranuloma venereum (LGV), a more invasive specimens are best collected by scraping the mucosa. sexually transmitted disease.  in addition, depending upon the site of involvement, blood, o LGV is characterized by transient papules on the external respiratory secretions, sputum, lung and other tissues can be genitalia, followed in 1 to 2 months by painful swelling of collected. inguinal and perirectal lymph nodes.  In case of LGV (Lymphogranuloma venereum), pus from the bubo should be collected. These specimens are processed  Trachoma: as: (See next page) o Chlamydia trachomatis, serotypes A, B, Ba, and C, cause a chronic keratoconjunctivitis that often results in A. LIGHT MICROSCOPY blindness.  Chlamydia trachomatis infections of conjunctiva, urethra and o Trachoma is transmitted by personal contact, for example, cervix may be diagnosed by demonstrating typical reniform from eye to eye via droplets, by contaminated surfaces inclusion bodies surrounding the nucleus after Giemsa staining, touched by hands and conveyed to the eye, or by flies. Macchiavello, Castaneda Methods. o Because of persistent or repeated infection over several  As the inclusion bodies possess a glycogen matrix, therefore, years, the inflammatory response with attendant scarring they may be stained with iodine solution also. leads to permanent opacities of the cornea and distortion of  This method has low specificity and sensitivity. eyelids.  BS MLS 3A DEANNASAURSAAAAAAA B. IMMUNOFLUORESCENCE  Species or genus-specific antigens are stained with fluorescent labelled antibodies (usually monoclonal antibodies).  The method has 90% sensitivity and 95% specificity.  And quick method. Within one hour, the results can be obtained. C. ELISA FOR THE DETECTION OF CHLAMYDIAL ANTIGENS  soluble genus-specific antigen captured by antibody attached to a solid surface like plastic body or microtiter well are detected.  the detection is based on the enzyme-labeled detector system and a chromogenic substrate.  The sensitivity and specificity of ELISA is similar to that of Immunofluorescence. D. DNA PROBES  DNA hybridization can be used for the direct detection of Chlamydia trachomatis in conjunctival and cervical smears. E. CHEMILUMINISCENCE ASSAY  Acridium-ester-labelled stranded DNA Probe is used which is specifically complementary to RNA of Chlamydia trachomatis or others based on need.  The DNA-RNA hybrid is detected in luminometer which measures light emitted by the acridium ester label.  Sensitivity and specificity is 95%. F. PCR  The omp1 gene and the 16s rRNA gene can be amplified and detected by PCR.  detecting these DNA has specificity up to 95% for identification up to species level and strains.