Spirochetes PDF

Spirochetes  Spirochetes Responsible for a wide variety of clinical conditions Order: Spirochaetales 5 genera with only 3 genera medically important) Divided into two families: Spirochaetaceae Borrelia Treponema Leptospiraceae Leptospira...

Spirochetes  Spirochetes Responsible for a wide variety of clinical conditions Order: Spirochaetales 5 genera with only 3 genera medically important) Divided into two families: Spirochaetaceae Borrelia Treponema Leptospiraceae Leptospira Spiro = Coiled Chaete = Hair General Characteristics Gram negative cell wall Long, slender, flexuous body with a size of: Wide = 0.1 - 0.5 um Length = 5 - 20 um Helically shaped and unicellular bacteria Flexible cell wall surrounded by several fibrils known as axial filaments/periplasmic flagella/endoflagella (gives Corkscrew-like motility) covered by an outer sheath attached to insertion disk May be free living or survive in animal and human host MANNER OF REPRODUCTION Treponema - Transverse fission Borrelia & Leptospira – Binary fission  Leptospira General Characteristics Both free-living and parasitic organism Cannot be readily stained using Giemsa or Tightly coiled Gram stain, but we can use silver staining to Thin visualize it. Flexible cell wall/ spirochetes If the bacteria is unstained it will not be visible Unique to these spirochetes is the presence of in bright field microscopy, but it can be seen in hooked ends in one or both ends of bacteria special microscope such as Dark field giving it a hook like appearance or question microscopy or Phase-contrast microscopy mark like appearance Bright field microscopy- used in the Motile with 2 periplasmic flagella or axial laboratory filaments, and 3-5 insertion discs. L. interrogans Agent of Human and Animal Leptospirosis Pathogenic strain that causes leptospirosis Common serotypes: L. interrogans serovar Icterohaemorrhagiae- (most common) L. interrogans serovar canicola L. interrogans serovar australis  Epidemiology Leptospirosis is the most widespread Leptospirosis is primarily associated with zoonotic disease found globally, although occupational and recreational exposure such discovered nearly hundred years ago it is still as during swimming, canoeing, farming or considered as an important emerging hunting Working with rat- or rat-infested disease. facilities poses a threat of infection. The people at risk includes but not limited to The organism is carried by dogs, rats or other farmers, veterinarians and slaughter house rodents. Infected animals harbors the workers bacteria in their renal tubules, in high concentration without significant untoward The bacteria may infect most mammals effect. So this bacteria will be shed during including amphibians and fishes micturition or your urination Leptospira can survive in neutral or slightly alkaline water for months  Pathogenesis The mode of transmission for Immune Phase Leptospirosis is direct with body fluid or indirect contact with contaminated The most common signs and symptoms waters, infected urine or blood of includes uveitis or the inflammation of the infected animal eye, rashes, hepatic and renal damage The disease has an incubation phase of 10 The hallmark of Immune Phase is the to 12 days but the disease may begin 2 to manifestation of what you call “Aseptic 20 days after the initial infection Meningitis”, along with CSF pleocytosis The most common form of the disease is The severe form of the disease that affects called “Anicteric Leptospirosis minority is the Icteric Leptospirosis or also known as Weil’s Disease Septicemic Stage Preventing Leptospirosis To prevent infection, it is important to take precaution on possible contact with body fluids of infected animals: Wear protective clothing (water resistant boot and gloves) Cover skin lesions with waterproof dressings Don’t wade or swim in potentially contaminated water Wash or shower after potential exposure Clean your wounds Don’t touch sick or dead animals Consume clean drinking water Rodent elimination Drainage of contaminated water Proper wound management Proper hand hygiene  Treatment Wear protective clothing (water resistant boot and gloves) Cover skin lesions with waterproof dressings Don’t wade or swim in potentially contaminated water Wash or shower after potential exposure Clean your wounds Don’t touch sick or dead animals Consume clean drinking water Rodent elimination Drainage of contaminated water Proper wound management Proper hand hygiene Laboratory Diagnosis Although not routinely performed, Leptospira can be cultured and isolated by various body fluids such as blood, CSF and urine. SAMPLE COLLECTION First week / 10 days- Blood o CSF During the acute phase of the illness, which is the first week or the first 7 to 10 days, blood and CSF may yield a higher chance of recovery. Second week up to 30 days – urine However, beginning the second week of illness for up to 30 days, urine is the recommended specimen. Laboratory Diagnosis Although not routinely performed, Leptospira can be cultured and isolated by various body fluids such as blood, CSF and urine. MICROSCOPIC EXAMINATION Direct: Dark field Microscopy / Phase-contrast Not recommended Microscopy Microscopic Blood, CSF, and urine may be examined directly by Dark field Microscopy / Phasecontrast Microscopy analysis, however, is not recommended. You may also opt to do silver staining or silver impregnation to visualize the Leptospira using Since, only a small Brightfield Microscopy. Sodium Oxalate / Heparinized blood / Urine percentage of cases are successful in Blood collected must used sodium oxalate and heparin as their anticoagulants. visualizing the organisms. For urine, it must not be placed in any preservatives and must be process within one There is a high chance hour. of false positive if the specimen used is urine. CULTURE The definitive test for Leptospira is the isolation using culture Media: Fletcher’s, Stuart, EllinghausenMcCullough-Johnson-Harris (EMJH) Sodium Oxalate or heparinized blood / Urine (Diluted 1:10) Direct inoculation of 1-2 drops of freshly drawn blood to the media may be used but if the sample is urine, it must be diluted 30°C in the dark 6 to 8 weeks To minimize the effect of inhibitory substances, cultures must be incubated at 30°C for 6-8 weeks in the dark The cultures must be checked weekly for the signs of growth A drop taken from a positive culture is detected using microscopy - Serodiagnosis or Molecular techniques  SERODIAGNOSIS In most diagnostic laboratories, the diagnosis of Leptospira is done via SERODIAGNOSIS IgM is detected within 1 week after disease onset of disease IgG detected or appear a month after onset Antibodies may be detected using ELISA or Enzyme -linked Immunosorbent Assay Macroscopic slide agglutination test is a rapid test, and the GOLD STANDARD for testing  Borrelia General Characteristics Microaerophilic organisms with loosely coiled bodies Just like other spirochetes, it is highly flexible with 30-40 axial filaments and 2 insertion discs It measures roughly to 0.2-0.5 micrometers in terms of diameter/thickness and 3-20 micrometers in terms of length. In contrast to leptospira, Borrelia stain readily by Giemsa stain and can be viewed under bright field microscopy "All pathogenic species of Borrelia are arthropod-borne” B. burgdorferi Causes lyme disease B. Reccurentis & B. duttonnii Cause relapsing fever  Relapsing Fever Based on the word relapse: There is a repeat acute febrile episodes that will subside and then will recur again after sometime. Virulence Factors Complement suppressor :C4b binding protein and Factor H Antigenic Variation Epidemiology Transmitted via vector which is an arthropod ❖ B. recurrentis: Louse: Epidemic RF ❖ transmitted by body Louse named pediculus humanus(organism) B. dutonnii: Tick: Endemic Relapsing Fever (RF) Human disease often occurs in small clusters following exposure to vector while sleeping in rustic cabins, seldom occupied rural dwelling, or caves PATHOGENESIS Incubation phase: 2-15 days During this time a massive number of spirochetes are developing in the bloodstream during the entire phase of relapsing fever. Symptoms include fever, headache, and myalgia that may last for 4-10 days Findings include petechiae, abdominal tenderness, and conjuncthyroid effusion. As the host mount a specific immune response against organism, the Borrelia organism disappears or sequesters from the bloodstream into various organs, so there will be a febrile period followed by a reemergence of the bacteria with newly synthesized antigens, hence the relapse of the fever.  LABORATORY DIAGNOSIS The most common method of diagnosing Borrelia is through the use of peripheral blood smear. Up to 70% of cases has visible organism when the specimen is collected during the febrile stage of the disease. Borrelia can also be cultured using Kelly medium (nutritionally rich agar), however it is rarely attempted and the possible yield is low. However, if the cultured is being done it must be incubated in microaerophilic conditions. Serodiagnosis against bacteria is also possible but not reliable due to antigenic variation of the bacteria. TREATMENT ❖ Borrelia is susceptible to many antimicrobials however tetracycline is used because it also reduces the number of relapse. ❖ However, one must be wary of the treatment, since Jarish- Herxheimer reaction may occur. Which is characterized as transient immunological reaction due to sudden released bacterial antigens following antibiotic treatment. ❖ Accompanied by fever, chills, headache, myalgia, cutaneous lesions, and death. Lyme Disease Lyme disease is caused by B. burgdorferi Binds Urokinase plasminogen activator (uPA) and Plasminogen ❖ That activates the plasmin, a protease that facilitates further invasion of the tissue Binding Factor H ❖ Allows the complement evasion and suppression of the immune system Stimulate TNFa ❖ Which is a potent suppressor of the immune system.  EPIDEMIOLOGY Most common tick-borne disease in United States and Europe Named after Lyme, Connecticut where a high incidence occurred in 1975. Reservoir: Mouse and Deer and Domestic animals MOT: Tick Bite (Ixodes) The vector are one of the genus Ixodes, where it can harbor the bacteria at any stage of its development. So it must be denoted that ticks require at least 24 hours of attachment to the host before they can pass the disease. PATHOGENESIS There are 3 stages in Lyme disease. In the 1st stage, is the appearance of erythema chronicum migrans (ECM). So this is a red ring lesion with a central clearing. Other symptoms include fever, headache, and malaise. The 2nd stage is characterized by disseminated infection. So it will occur weeks to months after the infection. So it will result to lesions, migratory joint and bone pain, alarming neurologic and cardiac diseases, splenomegaly, and severe malaise. For the 3rd stage, it is characterized by neurologic disease, chronic arthritis, and acrodermatitis chronica atropohicans (ACA) which is a diffused rash that may last for years.  LABORATORY DIAGNOSIS For isolation of Lyme disease, Borrelia burgdorferi is readily visualized in tissue section following staining by Warthin-Starry silver stain. CULTURE: Laboratorians may also attempt to culture the bacteria using Kelly Medium. Culture must be incubated in 30-34 degrees C for up to 12 days in microaerophilic conditions. However the preferred method of diagnosis is by SERODIAGNOSIS. The earliest antibody is IgM against OSpC, membrane protein, and flagellar antigens, and also fibronectin binding protein. For your IgG, it develops slowly against OSp17, p39, and p58. So this are antibodies that may be detected using IFA, EIA, and Western Blot. So the antibodies may be detected first using IFA (Immunofluorescence) and EIA (Enzyme Immunoassay). If negative, no further testing is required. However, if one of the test is positive, it must be confirmed using Western Blot.  Treponema General Characteristics Just like other spirochetes, they are thin, spiral organism with flexuous movement They have a size range of 0.1-0.2 micrometer in terms of thickness and width/diameter. And 6-20 micrometer in length. In contrast to Leptospira and Borellia, the coiling of Treponema are described as having a regular and angular. So, it has 6 -10 actual filaments with 1 insertion disc. They have a graceful flexuous movement in liquid They can be visualized using a Darkfield Microscopy. However, they are very thin and hard to visualize in Brightfield Microscopy. They are cultivable anaerobically or under anaerobic media/conditions on artificial media. T. pallidum subsp. pallidum It causes the venereal Syphilis, T. pallidum subsp. pertenue It causes Yaws T. pallidum subsp. endemicum It causes the endemic syphilis/ Bejel T. carateum It causes Pinta T. pallidum subsp. pallidum VIRULENCE FACTORS ▪ It has the ability to cross placenta and intact mucous membranes ▪ It is also possible that it can change its antigens, allowing it to evade the immune system. (ANTIGENIC VARIATION) CLINICAL MANIFESTATION It is the cause or agent of Venereal Syphilis. That Is normally transmitted via direct sexual contact. However, transmission via nongenital contact with a lesion is also possible such as on the lips and transplacental transmission. It is also possible that it can be transmitted through blood transfusion. The course of syphilis is divided into three parts; Primary Syphilis Secondary Syphilis Tertiary Syphilis Primary Syphilis Characterized by primary lesions or ulcerations found in the genitals. They are also known as “Chancre”, that appears 10- 90 days after the infection. So, in contrast to Haemophilus, the chancre produced by this bacterium is hard, and firm. It is also painless. The lesion is swarming or has many treponemes and is extremely infectious Direct contact with these lesions will expose you to syphilis. The chancre in woman is not really obvious most of the time because commonly, it is found in the cervix or in the vaginal wall At this stage, there is no systemic sign observed. So, the only signs and symptoms that you will experience is the formation of the chancres  Secondary Syphilis Skin rash Condylomata Sores on palms After the organism reached a sufficient number, clinical manifestation of secondary syphilis becomes more apparent. This happens 2-12 weeks after the development of the primary lesions. Symptoms include fever, sore throat, lymphadenopathy, headache, lesions on the mucous membrane, condyloma latum (wart like lesions on the genitals, also a widespread rash) So all secondary lesions in this stage are infectious.  Latent Phase Disease becomes subclinical producing no symptoms but not entirely dormant since they can be diagnosed using serologic methods Relapses are common which happens at: Early latent stage: < 1 year after secondary lesions Late latent stage: > 1 year after appearance of secondary lesions Tertiary Syphilis Tissue destructive phase Occurs or appears 10-25 years after initial infection S/S: Gummas in skin, bones and liver; Neurosyphilis, Cardiovascular lesions Gummas – granulomatous lesion Cardiovascular lesions – cause aortitis or inflammation of aorta that will lead to aneurysms Patient is NON-INFECTIOUS Benign nodular tertiary syphilis Congenital Syphilis Transmission of Treponemes from mother to fetus Since it can cross the placenta Early onset: 2 years after baby was born, it resembles the tertiary syphilis causing congenital defects like: Bone and Tooth deformities Cranial 8 nerve deafness Neurosyphilis Laboratory Diagnosis Mostly relies on the use of serologic testing and microscopic analysis Because Treponema is NON- CULTURABLE, hard to stain using Giemsa and Gram stain Can be visualized using Darkfield microscopy SEROLOGICAL TESTS It is the primary method for diagnosing syphilis. There are 2 type serologic test, the non- treponemal tests, and treponemal tests.  NON-TREPONEMAL TESTS Detects the reaginic antibodies Venereal disease research Rapid plasma reagin (RPR) that develop against the lipids released from the damaged cells. Detects cardiolipin Uses charcoal, which enable the reading of flocculation Only used as a screening method (+) Flocculation macroscopically. for syphilis since it only detects antibodies against the lipid itself Uses serum as samples Uses serum as samples Positive results must undergo It is preferred in diagnosing Unheated Serum Reagin (USR) definitive tests which is the neurosyphilis, which utilizes treponemal tests. CSF laboratory test (VDRL) Toluidine red unheated serum test (TRUST) The most used non-treponemal tests are the: Enzyme-linked immunosorbent assay (ELISA)  TREPONEMAL TEST Detects the presence of antibodies against treponemal antigens. Definitive test The reagent used in this test came from rabbit testicular lesion. The most common treponemal tests are the: Treponema pallidum particle agglutination (TP-PA) ❖ Uses gelatin with treponemal antigens. Fluorescent treponemal antibody absorption (FTA-ABS) Uses fluorescence label antihuman globulin  T. pallidum subsp. pertenue Causative agent of Yaws Early lesions known as the mother yaws (seen in the early phase), it will develop Also known as frambesia tropica, pian, 3 weeks to months after the initial parangi, or buba infection. Followed by the development of daughter yaws, that came from the Resembles syphilis dissemination of the mother yaws, numerous treponemes are found in Most prevalent non-venereal this lesion. Most people after this stage treponematoses will enter the stage of latency. After latency, usually no other manifestation Characterized by having tropical will occur. However small percentage up infection of the skin bones and joints to 10% of people who contracted the Contracted during childhood with breaks diseases develop late yaws, which is in the skin allowing the entry of the characterized by irreversible bone, treponemes cartilage, and soft tissue damage T. pallidum subsp. endemicum Causative agent of Endemic Syphilis AKA Bejel. Causes Bejel or Endemic Syphilis ❖ Transmission: mouth-to-mouth contact indirect contact through sharing of domestic utensils or use of contaminated utensils ❖ Begins in childhood as a small mucous patch, often on the interior of the mouth, followed by the appearance of raised, eroding lesions on the limbs and trunk. ❖ Periostitis (inflammation) of the leg bones is commonly seen, and gummas of the nose and soft palate develop in later stages. Primary Lesions: Painless, white mucinous ulcers in the oral cavity and the nipples of breastfeeding women Secondary Lesions: Split papules or rashes Periostitis of long bones or hand, sabre tibia, juxta-articular nodules Late Stage: Most common symptoms: infection of the eye optic atrophy chorioretinitis Gangosa causes destruction of the nasopharyngeal cartilage  T. carateum Agent of pinta ❖Is a skin human disease which initially presents as a raised papule followed by generalized eruption of flat reddened areas and is followed by development of bluish discoloration and a subsequent loss of pigmentation [just like psoriasis] Transmitted by nonsexual skin contact THANK YOU!