Microbiology Facts from Previous Papers

·         INSPISSATION: Egg containing media

LJ Medium, Loeffler’s, Dorsett, Petroff’s Medium

·         TYNDALLISATION:

Sugar or Gelatin Containing media

·         To enhance pigment production of S.aureus: 5% Milk addition, Glycerol acetate + NA

·         Generally GP forms SPORE:

GPB- Bacillus, Clostridium

GPC- Sporosarcina

But GNB- Coxiella Burnetti

·         Exceptions to KOCH’s POSTULATES:

Gonococcus: 3^rd not followed

Lepra Bacilli: 2^nd not followed

·         CONCOMINANT IMMUNITY:

Schistosomes creating barrier against continuous reinfection aks NATURAL ZOOPROPHYLAXIS

·         PARATETIC HOST: host in which parasite merily remains viable without further development.

Aka TRANSPORT or MECHANICAL HOST.

Eg: fly

·         NIH Swab: T.saginata, E.vermicularis

·         AUTOINFECTION occurs in: T.solium, S.stercoralis, E.vermicularis SET

·         Human infection dead end in: T.spiralis, E.granulosus

·         Lecithinase produced by: B.cereus, Clostriddium, Vibrio

·         VMA ELISA is for detection of: Shigella, EIEC

·         Heaviest Egg: unfertilized egg of A.lumbricoides

Largest Egg: fasciolepsis buski

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·         Mycobacteria should be stained for acid-fast organisms, using either Ziehl-Neelsen stain or other like Kinyoun stain.

·         Calcofluor white binds to cellulose and chitin in the cell walls of fungi and fluoresces under long-wave length ultraviolet light. It may demonstrate morphology that is diagnostic of the species (eg, spherules with endospores in Coccidioides immitis infection).

Pneumocystis jiroveci cysts are identified morphologically in silver-stained specimens.

PAS is used to stain tissue sections when fungal infection is suspected.

After primary isolation of fungi, stains such as lactophenol cotton blue are used to distinguish fungal growth and to identify organisms by their morphology.

·         Coenorhabditis elegans (a nematode) was the FIRST MULTICELLULAR ORGANISM FOR WHICH THE ENTIRE GENOME WAS SEQUENCED.  ( First bacteria was H.influenzae.)

·         Robert Koch

Father of Medical Microbiology

Discoveries – Anthrax bacilli, tubercle bacilli, vibrio cholera

The result of Koch investigations into a bubonic plague epidemic in Calcutta in 1897 showed that rats were vectors of the disease. He also demonstrated that sleeping sickness is transmitted by the tsetse fly.

Other studies by Koch included leprosy, surra, rinderpest, Texas fever and malaria. Koch formulated systematic method for biological research (Koch's Postulates) which are still observed to this day.

In 1905 Koch won the Nobel Prize for Physiology or Medicine "For his investigation and

discoveries in relation to tuberculosis".

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• Detection of HIV RNA by nucleic acid amplification of the viral load is the best predictor of “progression to AIDS.” (Virology)

• HIV RNA PCR and sequencing of the amplified products may be used to monitor resistance to anti-HIV drugs. HIV patients with total CD4 lymphocyte counts of less than 200 CD4 cells/μL are susceptible to opportunistic infections such as those caused by Cryptococcus, Mycobacterium, and Pneumocystis. (Virology)

• Cyclospora is an ooidian parasite similar to Cryptosporidium. It causes food-borne diarrheal illness and has been associated with contaminated berries. (Parasitology)

• Giardia, a large flagellate with both cyst and trophozoite forms, is the most common parasitic disease in the United States. The disease is characterized by diarrhea, cramping, and fever. (Parasitology)

• Enterohemorrhagic E. coli causes bloody diarrhea and hemolytic uremic syndrome. The mode of action is production of Shiga-like toxin by E.coli. (Bacteriology)

• Vancomycin-resistant enterococci, methicillin-resistant Staphylococcus aureus (MRSA), and vancomycin-indeterminate S. aureus (VISA) are among the most feared nosocomial pathogens. A recently introduced antibiotic, quinapristin-delfapristin, effectively treats vancomycin resistant enterococci or the few vancomycin-indeterminate MRSAs that have occurred. (Bacteriology)

• Following an upsurge of tuberculosis in the early 1990s, cases of Mycobacterium tuberculosis infection have remained static. M. tuberculosis causes initial primary pulmonary infection as well as a chronic disease characterized by hemoptysis, loss of weight, and fever. (Bacteriology)

• Penicillin-resistant pneumococci (Streptococcus pneumoniae) may account for up to 40% of isolates of S. pneumoniae. Third- or fourth-generation cephalosporins may be used as alternative treatment as well as vancomycin and rifampin. (Bacteriology)

·         Ehrlichia, a recently emerging tick-borne pathogen, is transmitted by Ixodes scapularis, the same tick that transmits the Lyme disease bacterium.

Ehrlichia is also transmitted by the Lone Star tick, Amblyomma americanum. (Chlamydia, Rickettsia)

• Eastern equine encephalitis may be transmitted to humans by the bite of a mosquito, particularly in the northeastern United States. (Virology)

• Transfusion-associated babesiosis is a growing problem, particularly in the immunosuppressed or patients without a spleen. Tick-borne babesiosis caused by the same tick that transmits Lyme disease is an emerging infection. (Parasitology)

• Dengue fever, a viral illness transmitted by the Aedes mosquito, is prevalent in epidemic proportions in both the Caribbean and Southeast Asia. (Virology)

• There are five major classes of immunoglobulin: IgG, IgM, IgA, IgD, and IgE. These immunoglobulins are distinguished by differences in the C regions of each individual H chain. These differences are function-related.

• Peptidoglycans are unique to prokaryotic organisms. They consist of a glycan backbone of muramic acid and glucosamine as well as crosslinked peptides. The enzymes responsible for cross-linking (transpeptidases) are the targets for β-lactam antibiotics. (Physiology)

• Genetic exchange in microorganisms occurs by several mechanisms, including transformation, transduction, conjugation, and transposition.

These processes are the basis for gene cloning in microorganisms. (Physiology)

• Virulence factors in bacteria include adherence factors, invasins, capsules, endotoxin, and exotoxin. Such factors enable microorganisms to invade the host, cause disease, and resist host defense mechanisms.(Physiology)

• Sites of action of antimicrobial agents include cell-wall synthesis, cell membrane integrity, DNA replication, protein synthesis, DNA dependent RNA polymerase, and folic acid metabolism. (Physiology)

• Staphylococcus aureus expresses two types of superantigens: enterotoxin (responsible for staphylococcal food poisoning) and toxic shock toxin.

• Free radicals of oxygen (superoxides) kill anaerobic bacteria exposed to air. Superoxide dismutase is a potent bacterial antioxidant. The presence of peroxidases in bacteria is protective. (Physiology)

• Campylobacter and Helicobacter are both helical-shaped bacteria. Helicobacter is known to play a role in the pathogenesis of peptic ulcer disease, while Campylobacter causes a food-borne gastrointestinal illness, most commonly from undercooked meat. Both bacteria are susceptible to antibiotics such as tetracycline. Helicobacter may be treated with Pepto-Bismol, metronidazole, and amoxicillin. (Bacteriology)

• The agents of bovine spongiform encephalopathy (Mad Cow Disease), scrapies, and new-variant Creutzfeldt-Jakob disease in humans are prions or amyloid fibrils. Also included are prions that cause chronic wasting disease (CWD) in elk and deer, although these agents of CWD have not been shown to be transmissable to either cattle or humans. These self replicating proteins are resistant to heat and chemical agents. (Virology)

• Prior to 1999, West Nile virus, an arbovirus with serological crossreactivity to St. Louis encephalitis virus was not seen in the United States. However, during 1999 and 2000, a large number of birds were infected with West Nile virus, as well as a few humans, some of whom died. (Virology)

• The genotype of hepatitis C is important in predicting the response of this virus to therapy with interferon and ribavirin as well as the required length of treatment. (Virology.

·         Vascular invasion is characteristic of: mucormycosis, aspergillus

2) Inhalation of spores of which fungus is commonest: aspergillus

3) Only pathogenic yeast in man: C. neoformans

4) Sclerotic bodies with septate hyphae characteristic of: blastomycosis

5) Asteroid bodies are a characteristic feature of: sporothrix

6) Fungal infection is characterised by its capability to disseminate via lymphatics: sporothrix

7) Sporulation occurs in which phase: phase of decline

8) Prominent feature of paracoccidiomycosis: ulcerative granuloma of buccal and nasal mucosa

9) Characteristic feature of coccidiomycosis: spherule formation

10) Fungal infection which causes arthralgia / valley fever / desert rheumatism:
Coccidiomycosis

11) Thick walled barrelled shaped spores: arthrospores characteristic of coccidiodes immitis

12) Fungus which is urease positive: C. neoformans

13) Copper pennies appearance in tissue: paracoccidiodes

14) Most characteristic clinical feature of all cases of paracoccidiodomycosis: lymph node enlargement

Infectious Doses

·         Shigella, enterohemorrhagic Escherichia coli, Giardia lamblia, or Entamoeba, as few as 10–100

·         Salmonella: 10³–10⁶

·         Vibrio cholera: 10⁵–10⁸

·         ETEC, EIEC, and EAggEC: 10⁶ to 10¹⁰

·         Cryptosporidium: 50% infectious dose: ~132 oocysts in nonimmune individuals

·         B. anthracis, While an LD50 of 10,000 spores is a generally accepted number it has also been suggested that as few as one to three spores may be adequate to cause disease in some settings.

·         F.turarensis: as few as 10 organisms causing infection

Immunology

·         PRIMARY IMMUNODEFICIENCY’S:

1.       Antibody/B cell deficiencies [Bruton's X-linked agammaglobulinemia, AR agammaglobulinemia, CVID (ICOS and TACI deficiencies), HIM (AR), ICF syndrome]

2.       Cellular deficiencies (IFN-γ/IL-12 axis, APS type 1, defective NK function)

3.       Combined [SCID, Wiskott-Aldrich, DiGeorge, ataxia telangiectasia, HIM (AR), XLP syndrome, WHIM syndrome, caspase 8 deficiency]

4.       Complement

5.       Phagocytic (CGD, Chediak-Higashi syndrome, LAD, specific granule deficiency, cyclic neutropenia, X-linked neutropenia)

·         ASSAYS FOR CELLS:

T cells: Flow cytometry for CD3, CD4, CD8, CD45, T-cell receptor

B cells: CD19, CD20, CD21, Ig-associated

NK cells: CD16/CD56

·         B Cell-Signal-Transduction Complex: Ig-a;, Ig-l3,CD19, CD20, CD21

·         T-Cell Receptor (TCR): CD3 complex (gamma, delta, epsilon and zeta chain dimers)

·         IgG has highest Affinity.

IgM has highest Avidity.

·         ROUTINE HLA TYPING:

Done for HLA-A, HLA-B, HLA-DR

The micro cytotoxicity assay is used to identify class I MHC molecules. This test is performed using antisera against specific class I antigens, plus complement. If the test cell has class I antigen to match the antibody, then complement will be bound and the cell will be killed. The entry of the dye into the interior of the cell illustrates cell death.

The mixed lymphocyte assay is used to test reactions between donor and recipient cells.T cells are incubated with a B cell to check the response to class II (DR) antigens. The B cells act as the stimulator cell, and they are irradiated to prevent proliferation. The T cells respond because they recognize MHC molecules on the irradiated (stimulator) cellthat are different from their own MHC class II molecules. Proliferation is measured by uptake of titrated thymidine in cellular DNA.

·         Because the B cells divide rapidly in germinal centers of the lymph nodes, there are many opportunities for mutations. Although these mutations can be detrimental to individual B cells (these cells are then eliminated), mutations that increase antibody affinity are preferentially selected, and mutations thus playa role in affinity maturation of B-cell clones. Somatic mutation, however, plays essentially no role in T-cell receptor diversity.

·         TAP-2 deficiency (also known as bare lymphocyte syndrome I) is caused by defects in the transporter associated with the antigen presentation (TAP; either TAP1 or TAP2) system. These defects ultimately impair the loading of peptide fragments into nascent MHC class I molecules in all nucleated cells and reduce the number of MHC class I molecules that successfully reach the cell surface. This reduced MHC I expression decreases the number of functional CD8+ T cells and can also affect the functions of NK cells monitoring MHC class I expression on body cells (although the NK cells appear not to attack uninfected host cells). Likewise, defects inhibiting expression of MHC class II molecules reduce the number of functional CD4+ T cells.

·         Some IgA-deficient patients with anti-IgA antibodies are at increased risk of a severe reaction to trace IgA from IVIg and from blood transfusions (washed packed cells should be given if transfusion is needed).

·         Persons with IgG deficiencies, As a rule are not susceptible to viral infections, However, IgG-deficient patients are susceptible to polioviruses (and should not receive live virus vaccine) and to hepatitis B and C.

In patients with XLA (but not CVID or HIM), a chronic meningoencephalitis, which is ultimately fatal, can develop with enteroviruses (echovirus or coxsackie) infection.

·         Different types of SCID:

1.       ADA deficiency

2.       PNP deficiency

3.       RAG1, RAG2 (Omenn syndrome)

4.       Defective cytokine signaling

5.       X-linked (common gamma chain) this one is MC

6.       Autosomal recessive (IL2, IL7 receptor)

·         Idiopathic CD4 lymphocytopenia (Nezelof syndrome) isT cell immunodeficiency; just like DiGeorge syndrome without the associated congenital anomalies; resembles HIV (failure to thrive, candidiasis, anemia, thrombocytopenia, cancer).

·         Chronic mucocutaneous candidiasis (autoimmune polyglandular syndrome type I) is T cell immunodeficiency; superficial candidiasis (not systemic) associated with single or multiple endocrinopathies, iron deficiency, and anergy; molecular defect in AIRE

·         Hyperimmunoglobulin E is a/w Uncertain immunodeficiency, with increased serum levels of IgE (up to 10 times normal); recurrent infections of the skin and sinopulmonary tract with S. aureus, H. influenzae, and Aspergillus; coarse facial features; delayed shedding of primary teeth; and chronic eczematous rashes.

·         CVID; chronic mucocutaneous candidiasis and hyperimmunoglobulin E are disorders compatible with living to adulthood; the other T-cell or combined immunodeficiencies are severe and generally present early in life; without bone marrow transplantation, they are generally fatal.

·         Hyper IgM Syndrome: X linked mutation in CD40 ligand on T cells. So no class switching. IgM level 10 times normal. Deficiency of IgA, IgG, IgE. Fails to make Germinal Center. Recurrent Respiratory Infections.

·         Anaphylactoid reaction is clinically similar to anaphylaxis but is not IgE-mediated. Agents such as radiocontrast media, NSAIDs, muscle relaxants, and paclitaxel cause direct stimulation of mast cells and basophils. Anaphylactoid reactions can often be prevented with glucocorticoids and antihistamines.

·         If the C4 level is normal during an episode of angioedema, there is no problem with C1INH because C4 is used up in this process. If the C4 level is low or if a person is seen in an asymptomatic period, C1INH level and functional activity should be measured (to determine if it is HAE I or II), as should the C1q level.

C1q levels are normal in hereditary angioedema and decreased in acquired C1INH deficiency.

C2 is reduced during acute episodes.

 Acquired C1INH is associated with malignancy, particularly B-cell lymphomas.

·         C1INH deficiency is treated with attenuated androgenic steroids, which increase the production of C1INH. This is effective in patients with deficient production, deficient activity, and increased catabolism of C1INH. Epinephrine may not work in a crisis, and a tracheostomy is indicated for laryngeal edema. Antifibrinolytics (epsilon-aminocaproic acid or tranexamic acid) may be helpful.

·         DIAGNOSIS OF MASTOCYTOSIS:

One major diagnostic criterion + one minor criterion or three minor criteria

Major criteria: Bone marrow biopsy and aspirate with dense infiltrates of mast cells

Minor criteria:

1         Spindle-shaped mast cells

2         Detection of 816 c-kit mutation

3         Flow cytometry with CD2, CD25, and CD117 expression

4         Serum tryptase >20 µg/mL

·         TRANSPLANT MATCH :

Although matching for intrafamilial transplants of all types is performed, nonfamilial cardiac, lung, and liver grafts are not MHC-matched because other factors such as size, location, and availability limit the transplants much more. Kidney transplantation, for which there is the potential for living related and unrelated donors, allows for matching. Bone marrow transplants must be matched, whereas matching in liver transplants may actually decrease survival.

·         Graft-versus-host disease is an immune response of the donor T cells against the recipient usually 6 or more days after transplant. It is a problem only when transplanting hematopoietic tissue (bone marrow, nonirradiated blood transfusions).

·         Pneumococcal polysaccharide vaccine (PPV) protect against 23 pneumococcal bacteria.

·         Of the many methods available for antigen and antibody detection, LA, ELISA, EMIT, CIE, and COA are the most widely used.

Latex agglutination (LA) employs latex polystyrene particles sensitized by either antibody or antigen. LA is more sensitive than CIE and COA but slightly less sensitive than either RIA or EIA. LA has been used to detect Haemophilusinfluenzae, Neisseria meningitidis, and Streptococcus pneumoniae antigens in cerebrospinal fluid. LA has also been used for detection of cryptococcal antigen. Most recently, LA has been widely used for rapid detection of group A streptococcal antigen directly from the pharynx. The test is rapid (5 minutes), sensitive (approximately 90%), and specific (99%).

Coagglutination (COA), also an agglutination test, is slightly less sensitive than LA but less susceptible to changes in environment (e.g., temperature). Most strains of coagulase-positive staphylococci have protein A in their cell wall. Protein A binds the Fc fragment of microbial antigens in body fluids. COA has also been used to rapidly type or group bacterial isolates.

Enzyme immunoassays (EIAs) can be either homogeneous (EMIT) or heterogeneous (ELISA). EMIT has been used primarily for assays of low-molecular-weight drugs. Its primary use in microbiology has been for assays of aminoglycoside antibiotics. EIAs vary as to the solid support used. A variety of supports can be used, such as polystyrene microdilution plates, paddles, plastic beads, and tubes. The number of layers in the antibody-antigen sandwich varies; usually as additional layers are added, detection sensitivity is increased. The two most common enzymes are horseradish peroxidase (HRP) and alkaline phosphatase (AP). Beta galactosidase has also been employed. Orthophenylene diamine is the most common substrate for HRP and p-nitrophenyl phosphate for AP. Because EIAs are usually read in the visible color range, the tests can be read qualitatively by eye or quantitatively by machine.

Counterimmunoelectrophoresis (CIE) was originally used for "Australia antigen" (HBsAg) but was soon replaced by RIA. For a decade, CIE was used to detect antigens in body fluids, CIE is not an easy technique. Its success depends on the control of many variables, including solid support, voltage, current, buffer, affinity and avidity of antibodies, charge on the antigen, and time of electrophoresing.

• While acute stress tends to suppress the immune system, chronic stress tends to enhance the immune system, especially over time.

• If the immune system is weakened so that thelatent virus becomes more active, the antibody titers to the virus will increase. Relaxation training produces decreased antibody titers to latent viruses by generating an increase in natural killer cell activity.

• Consumption of alcohol has been shown to affect the immune system, apparently in relation to dose. Macrophages are inhibited and move more slowly toward chemical signals released by the body. Proliferation and cytotoxicity of T cells are also inhibited. Increased doses of alcohol can damage certain bodily tissues, such as the thymus. However, no inhibition of production of antibodies has been observed.

·         The brain and the endocrine and immune systems communicate with and influence each other. Studies of the hypothalamus have demonstrated that stimulation of the anterior hypothalamus enhances immune responsiveness, and stimulation of the posterior lobe inhibits immune reactivity. Removal of the thymus gland will result in the atrophy of these hypothalamic neurons. Also, the limbic system contains high concentrations of neuropeptide receptors, as do the lining of the intestines and lymphocytes. It has also been found that immune cells produce the same neuropeptides that are produced in the brain. Some of these peptide hormones that are produced in the brain stimulate T cells to produce lymphokines, such as interleukins and interferon.

·         R. Ader and N. Cohen discovered that the immune system could be conditioned by neutral  stimuli.

Robert Ader, also discovered that the immune system can be suppressed through classical conditioning.

·         A battery of immunologic assessments found that the caregivers had suppressed immune systems; cellular immune system control of latent viruses was poorer than that of a matched control group, the percentage of T lymphocytes was lower, and the helper/suppressor ratio was smaller.

·         99% Proteins are antigens. Cardiolipin (seen in SLE) is the ONLY FAT that is an antigen.

·         Electrophoresis of human serum proteins identifies five distinct types: albumin, 1-proteins, 2-proteins, -proteins, and globulins.

Many human diseases can be diagnosed, at least in part, on the basis of abnormal electrophoresis profiles. For example, absence of the second peak) is compatible with a diagnosis of Alpha 1-antitrypsin deficiency in symptomatic persons. A sharp and high peak indicates the presence of a monoclonal gammopathy, such as multiple myeloma; on the other hand, a peak that is diffusely elevated points to polyclonal hypergammaglobulinemia. Complete absence of the peak is associated with Swiss-type agammaglobulinemia.

• The complement-fixation (CF) test is a two-stage test. The first stage involves the union of antigen with its specific antibody, followed by the fixation of complement to the antigen-antibody structure. In order to determine whether complement has been "fixed," an indicator system must be employed to determine the presence of free complement. Free complement binds to the complexes formed when red blood cells (RBCs) are mixed with anti-RBC antibody; this binding causes lysis of the cells. Complement that has been "fixed" before addition of red blood cells and anti-RBC antibody cannot cause lysis.
• A DNA probe is available. It is an 125I probe for the 16S ribosomal RNA of M. pneumoniae.
• Falsely positive ELISAs and Western blots is an influenza vaccination within the past few months.

• COMPLEMENT AND DISEASE:

Patients with deficiencies of the middle complement components (C3 and C5) are at risk for recurrent pyogenic infections,. Individuals with C3 deficiency have recurrent serious pyogenic bacterial infections that can be fatal. The absence of C3 leads to the inability to generate the opsonin, C3b, which, when deposited on the surface of the bacteria, promotes phagocytosis. Isolated C3 deficiency typically presents at a very early age, most often shortly after birth.

Individuals with C2 deficiency have a predisposition for immune complex disease such as systemic lupus erythematosus.

while those lacking terminal complement components (C6, C7, or C8, but not C9) are prone to developing recurrent infections with Neisseria species.

A deficiency of decay-accelerating factor (DAF), which breaks down the C3 convertase complex, is seen in paroxysmal nocturnal hemoglobinuria (PNH), a disorder that is characterized by recurrent episodes of hemolysis of red cells because of the excessive intravascular activation of complement.

Deficiencies of C1 esterase inhibitor result in recurrent angioedema, which refers to episodic nonpitting edema of soft tissue, such as the face. Severe abdominal pain and cramps, occasionally accompanied by vomiting, may be caused by edema of the gastrointestinal tract. To understand how a deficiency of C1 inhibitor can cause vascularly produced edema (angioedema), note that not only does C1 inhibitor inactivate C1, but it also inhibits other pathways, such as the conversion of prekallikrein to kallikrein and kininogen to bradykinin. A deficiency of C1 inhibitor also leads to excess production of C2, a product of C2 called C2 kinin, and bradykinin. It is the uncontrolled activation of bradykinin that produces the angioedema, as bradykinin increases vascular permeability, stimulates smooth muscle contraction, dilates blood vessels, and causes pain.

·         CLASS-SWITCHING: Changes in the constant-region germline, to change isotypes (from IgM to IgG, etc.)

Antigen binding to a BCR (IgM) causes switching from IgM or IgD ------> IgG, IgA or IgE.

The V region remains the same, such that the cell retains its same specificity, but the constant regions change to a different class.

-          SWITCH SITES: This occurs by a process called switch recombination, in which all constant-region genes (DNA) are deleted in progeny, except the ones being expressed.

Switch sites are located in the introns, proximal (5') to each CH locus.

They contain tandem repeats that are highly conserved across species.

-          REVERSIBILITY: Switching is for the most part irreversible, because intervening C regions are deleted. However, a C-region that remains downstream remains intact and can thus still be switched to.

-          Double Class-Switching is possible with help of IL-4, which induces class two class switches in a row in the process of making IgE:

mu () ------> gamma-1 (gamma1)

gamma-1 (gamma1) ------> epsilon ()

·         T-cell markers (e.g., CD4 and CD8) have functional significance, whereas B-cell markers (e.g., CD19 and CD20) are primarily of maturational significance.

·         Opsonization: Facilitating phagocytosis. Neutrophils have lots of opsonic receptors.

Neutrophils have Fc-Receptors, to recognize the Fc (common) portion of an IgG molecule bound to antigen. This extra affinity facilitates phagocytosis of the antigenic material.

Neutrophils have CR1, CR2, CR3 receptors, all of which bind C3b, a Complement degradation product. Thus, the complement pathway facilitates opsonization of antigen.

·         AGRETOPE: The region of the antigen that binds to the MHC complex. A given MHC molecule can bind a variety of different antigens.

·         EPITOPE: The region of the antigen that binds to the T-Cell Receptor (TCR).

·         IDIOTOPE: The individual antigenic determinant region of an immunoglobulin.

The idiotope may or may not be the same as the antigen-binding site, or it may overlap with it.

Each antibody has multiple idiotopes within it -- those regions capable of acting as antigen.

·         IDIOTYPE: The sum of the idiotopes of an immunoglobulin.

Monoclonal antibodies, all with same structure, all have the same idiotypes.

IDIOTYPIC DETERMINANTS: Common idiotopes that tend to show up in immunoglobulins even if they are polyclonal. This is due to the common germline structure of immunoglobulins.

·         ISOTYPE: Constant-region antigenic determinants that are species-specific. They are contained with the constant region of an Ig and are thus endemic to the species.

·         ALLOTYPES: Antigenic determinants that are inherited, and thus represent allelic variation within the constant region.

Only some of the constant region domains have allotypes (i.e. multiple alleles). Others do not, at least not known.

·         ANTIGENIC SHIFT: Making major variations to the viral protein coat, by means of Natural Selection and mutation. This makes it difficult to continually defend against the virus.

ANTIGENIC DRIFT: Minor variations to the viral protein coat. graDually, due to point mutation

·         MITOGEN: Substances that are non-specific polyclonal activators of lymphocytes. They induce polyclonal lymphocyte proliferation.

Lectins: Polyclonal activators that bind to common sugar-residues found on many cells. They can induce agglutination and then proliferation of lymphocytes.

CONCANAVALIN-A (ConA): A polyclonal T-Cell mitogen.

PHYTOHEMAGGLUTININ (PHA): A polyclonal T-Cell mitogen.

Pokeweed Mitogen: A polyclonal activator of both B and T cells.

·         PAPAIN: Protease cuts right above the joining region, generating three parts.

PEPSIN: Protease cuts right below the joining region, generating single main fragment

Parasitology

·         Nutrient poor agar with E. coli overlay provides nutrient for the growth of acanthamoeba.

Mycology

·         C. albicans is part of the normal flora of the gastrointestinal tract, mouth, and genital surfaces. Notwithstanding, C. albicans causes severe disease particularly in those patients with compromised immunity. It is generally thought that when C. albicans is unable to adhere to mucosa it is nonpathogenic and that production of germ tubes or hyphae plays a major role in colonization and infection of the mucosal epithelial cells by allowing direct penetration of these cells with specific hydrolytic enzymes. While other mutations such as temperature intolerance, metabolic alterations, and structural substitutions may affect the ability of Candida to survive, these changes would not affect adherence.

Virology

·         St. Louis encephalitis virus is spread by mosquitoes and causes a severe encephalitis with a near 10% fatality rate. Colorado tick fever, spread by the wood tick, causes fever, headache, retro-orbital pain, and severe myalgia. Fever and jaundice characterize yellow fever, a life-threatening disease spread by mosquitoes. Following fever, headache, myalgias, and photophobia, the symptoms progress to the liver, kidney, and heart. Mortality rate is high. Dengue fever shares the same mosquito vector as yellow fever. Classic dengue fever (breakbone fever) includes flu-like symptoms. Severe muscle and joint (breakbone) pain occurs. Coxsackievirus is spread by the fecal-oral route.

Bacteriology

·         An understanding of normal, or indigenous, microflora is essential in order to appreciate the abnormal. Usually, anatomic sites contiguous to mucous membranes are not sterile and have a characteristic normal flora.

The skin flora differs as a function of location. Skin adjacent to mucous membranes may share some of the normal flora of the gastrointestinal system. Overall, the predominant bacteria on the skin surface are S. epidermidis and Propionibacterium, an anaerobic diphtheroid.

The gastrointestinal tract is sterile at birth and soon develops a characteristic flora as a function of diet. In the adult, anaerobes such as B. fragilis and Bifidobacterium may outnumber coliforms and enterococci by a ratio of 1000:1. The colon contains 1011 to 1012 bacteria per gram of feces.

The mouth is part of the gastrointestinal tract, but its indigenous flora shows some distinct differences. While anaerobes are present in large numbers, particularly in the gingival crevice, the eruption of teeth at 6 to 9 months of age leads to colonization by organisms such as Streptococcus mutans and S. sanguis, both alpha-hemolytic streptococci. An edentulous person loses alpha-hemolytic streptococci as normal flora.

 Soon after birth, the vagina becomes colonized by lactobacilli. As the female matures, lactobacilli may still be predominant, but anaerobic cocci, diphtheroids, and anaerobic Gram-negative rods also are found as part of the indigenous flora. Changes in the chemical or microbiologic ecology of the vagina can have marked effects on normal flora and may promote infection such as vaginitis or vaginosis.

·         Two human forms of ehrlichiosis can occur:

human monocytic ehrlichiosis (HME), caused by E. chaffeensis; and

       human granulocytic ehrlichiosis (HGE), caused by an Anaplasma phagocytophilum.

Ehrlichiosis was previously recognized only as a veterinary pathogen. HME infection is transmitted by the brown dog tick and A. americanum. HGE infection is transmitted by I. scapularis, the same tick that transmits Lyme disease. Both infections cause fever and leukopenia. A rash rarely occurs. E. chaffeensis infects monocytes, and HGE infects granulocytes; both organisms produce inclusion bodies called morulae.

·         Listeria is a common inhabitant of farm animals and can be readily isolated from silage, hay, and barnyard soil. Humans at the extremes of age are most susceptible to Listeria infection but only recently has food been implicated as a vehicle. In the outbreak in Nova Scotia, it is likely that the cabbage used for the coleslaw was fertilized with animal droppings and not properly washed prior to consumption. Major Listeria outbreaks associated with cheese have been seen in the United States and most likely have originated from contaminated milk.

·         Adenosine triphosphate (ATP) is believed to be generated at three reaction points in the electron transport chain: the reductions of flavoprotein, cytochrome b, and cytochrome c. This phenomenon, demonstrated in experiments with mammalian mitochondria, can be expressed in terms of the relationship between the moles of ATP generated for each atom of oxygen consumed—the P/O ratio. In mammalian cells, the P/O ratio is 3; that is, there are three segments in the electron transfer chain in which there is a relatively large free energy drop. In bacteria, however, there appears to be only one or two of these segments. Loss of these phosphorylation sites as well as reactions that bypass these sites of ATP synthesis account for the lower P/O ratio in bacteria. Some bacteria, such as Mycobacterium phlei, have P/O ratios of 3.

·         Penicillin-resistant, non--lactamase-producing, vancomycin-resistant, Gram-positive cocci are most likely Enterococcus faecium. There are a variety of mechanisms for vancomycin resistance in E. faecium and they have been termed Van A, B, or C. These isolates have become one of the most feared nosocomial pathogens in the hospital environment. Unfortunately, no approved antibiotics can successfully treat vancomycin-resistant enterococci (VRE), only some experimental antibiotics such as Synercid.

·         Wound exudates smell bad owing to production of organic acids by such anaerobes as B. fragilis. Black exudates or a black pigment (heme) in the isolated colony is usually a characteristic of Bacteroides (Porphyromonas) melaninogenicus, not B. fragilis.

·         The structural integrity of the beta-lactam ring in penicillins is essential for their antimicrobial activity. Many resistant strains of staphylococci produce an enzyme, penicillinase, that cleaves the beta-lactam ring at the carbon-nitrogen bond. Other organisms, including certain coliform bacteria, produce an amidase enzyme that inactivates penicillin by disrupting the bond between the radical and nitrogen in the free amino group.

·         6-Phosphogluconic acid is a characteristic metabolic intermediate in the pentose-phosphate metabolic pathway. This pathway is used by heterolactic fermenters such as Leuconostoc, the organism responsible for the fermentation of cabbage in the production of sauerkraut. Leuconostoc is a Gram-positive bacterium with a dextran capsule.

·         Dipicolinic acid, formed in the synthesis of diaminopimelate (DAP), is a prominent component of bacterial spores but is not found in vegetative cells or eukaryotic appendages or fimbrial structures. The calcium salt of dipicolinic acid apparently plays an important role in stabilizing spore proteins, but its mechanism of action is unknown. Dipicolinic acid synthetase is an enzyme unique to bacterial spores.

·         ADP-ribosylates G_s = Turn the On, On

Vibrio

ETEC

·         ADP-ribosylates G_i = Turns the Off, Off

Bordetella pertussis

·         Ribosylation of EF₂

Pseudomonas

Diptheria

·         Common bacteria in the back of the throat

o  Strep. Pyogenes

o  S. pneumonia - encapsulated

o  H. influenza - encapsulated

o  Neisseria Cataralis - encapsulated

§  These bacteria can live in the back of throat because they contain IgA Protease to protect against IgA activity

§  Therefore, they are the MCC of sinusitis, otitis, bronchitis, and pneumonia

·         Enterococcus is the only nitrite negative UTI.

·         Cultures of Actinomyces grow as white masses with a domed surface, which is called a “molar tooth” appearance.

·         Nocardiae are aerobic and acid-fast, in contrast to Actinomyces species, which are strict anaerobes and not acid-fast.

Actinomyces israelli is treated with penicillin and Nocardia asteroides is sensitive to trimethoprim-sulfamethoxazole.

·         L. monocytogenes is a short, gram-positive, non-spore-forming bacillus that can produce neonatal disease or can result in stillbirth. Characteristics that are unique to Listeria include a tumbling motility on hanging drop and an umbrella-shaped motility pattern when a specimen is stabbed into a test tube agar slant.

Listeria may be mistaken for diphtheroids on Gram stain; both are gram-positive bacilli. Listerial infection often occurs in immunocompromised patients. Risk factors include cirrhosis, neoplastic disease, renal failure, pregnancy, chronic steroid therapy, and extremes of age (i.e., very young and elderly).

·         Bartonella infections are also characterized by proliferations of blood vessels. Examples of Bartonella include B. quintana, B. henselae, and B. bacilliformis,

B.bacilliformis, the causative agent of Oroya fever.

B. quintana is spread by the human body louse and is the causative agent of trench fever (seen in the trenches of World War I) and bacillary angiomatosis. This latter term refers to a lesion seen in patients with AIDS consisting of a lobular proliferation of capillaries with abundant leukocytoclastic debris.

 B. henselae is the causative agent of cat-scratch fever. Histologically, this disease is characterized by the formation of stellate microabscesses with necrotizing granulomas.

·         Persons with acute and chronic alcoholism become infected with Klebsiella pneumoniae, most often by aspiration. K. pneumoniae pneumonia often localizes in the upper lobe, sometimes the minor fissures bows downward, reflecting the bogginess of the upper lobe involved with this infection.

·         The process of transduction involves the transfer of a portion of DNA from one bacterium to the chromosome of another bacterium by means of a viral infection.

Conjugation is the transfer of a so-called male chromosomal DNA to the DNA of an acceptor, or female, bacterial cell.

Colinearity defines the relationship between genes and proteins in that the sequence of amino acids in proteins is a result of the sequence of base triplets in template genes.

 Recombination is simply the exchange of sequences between two molecules of DNA.

 Transformation results when exogenous DNA fragments are incorporated into the chromosome of another organism, as in the transformation of pneumococcal bacteria that led Avery and McLeod to recognize the genetic significance of DNA.

·         ENTEROBACTERIACEAE ANTIGENIC MARKERS:

1.       O-Antigen: Carbohydrate in the cell wall.

2.       K-Antigen: Capsular antigen, when present.

3.       H-Antigen: Flagellar antigen, when present: Campylobacter, Helicobacter, Escherichia.

4.       F-Antigen: Pili antigen.

·         ENTEROTOXINS A, B, C1, C2, D: Affects the GI-tract in S. AURES food poisoning.

They are heat stable -- heating (or reheating) food does not denature the toxin.

·         Virulence of S.saprophyticus:

Urease: Enable survival in urinary tract.

Tropic for urinary transitional epithelium.

Lysostaphin is lethal to S. AURES and other bugs.

·         B. anthracis species is identified by a String of Pearls formation in penicillin. The colonies are susceptible to penicillin, whereas other Bacillus species are not.

B. Cereus is penicillin-resistant. They will form chains of rods rather than string of pearls in penicillin suspension.

BACILLUS SUBTILIS at Risk: Heroin preparations can be contaminated with it.

·         FRANCISELLA TULARENSIS: CYSTEINE-AGAR must be used.

·         In Legionella, Dieterle Silver Stain shows rod shaped bacteria within macrophages. NO STAIN with Gram Stain, Acid-Fact Stain, or H&E. CYSTEINE is absolutely required for growth.

·         Y. pestis, Bacteria reproduce in flea to the point where they clog the proventriculus. Human bite ------> inoculation of bacteria into host.

·         Pyoderma of Streptococci:

IMPETIGO: Cutaneous lesions in kids, usually on face. Very similar to the Staph bullous lesions, except that these are vesicular lesions because these are actually full of bugs.

ERYSIPELAS: Cutaneous lesions in adults. Painful. Again contains organisms.

·         Todd-Hewitt Broth: Done when few organisms were recovered.

Strep: Granular precipitates on sides and bottom of tube.

Staph: Diffuse turbidity.

·         CAMP TEST: Mix normally non-hemolytic Staph Aureus with specimen, and hemolytic zone is augmented. Staph Aureus is normally only hemolytic at 4C.

·         ENTEROCOCCUS FAECALIS (GROUP D)

Virulence: LIPOTECHOIC acid, very lipid rich, leads to gram-variable appearance.

Treatment: Penicillin resistant, strongly, due to altered Penicillin-binding proteins.

Also have acquired vancomycin and gentamycin resistance.

·         Vincent's Disease can be confused with diphtheria.

 Vincent's Disease is pseudomembranous pharyngitis, and is caused by poor oral hygiene.

Vincent's will show halitosis and signs of gingivitis (swelling, bleeding).Stain with Vincent's disease will show a whole mess of bacteria -- both Gram (+) and Gram (-), while Diphtheria will show only slender Gram (+) rods.

·         P. Vulgaris: Positive for metabolizing tryptophan ------> indole

Ampicillin-resistant. Use aminoglycoside and then wait for sensitivity tests.

P. Mirabilis: Negative for metabolizing tryptophan ------> indole

Treated with Ampicillin

·         NEISSERIA MENINGIDITIS (MENINGEOCOCCUS)

Type-B is the most virulent because it is not immunogenic -- we cannot make protective antibodies against it.It is made out of alpha-2,8-n-acetylneuraminic acid. It is rapidly degraded and thus not immunogenic.

LPS: Induces the Schwartzman Reaction. Causes the rash and can lead to necrosis.

·         BRUCELLOSIS:

Brucella Agar = Selective Agar containing Erythritol, which the bugs subsist on.

Erythritol is found in reproductive tract of cattle, hence these bugs cause abortion in cattle. Humans don't have erythritol so no abortion happens.

·         BORRELIA BURGDORFERI:

Enriched Selected Kelly's Medium. Enriched with fatty acids, but it's still difficult to grow.

·         The 16S rRNA of each species of bacteria has stable (conserved) portions of the sequence. Many copies are present in each organism. Labeled probes specific for the 16S rRNA of a species are added, and the amount of label on the double-stranded hybrid is measured. This technique is widely used for the rapid identification of many organisms. Examples include the most common and important Mycobacterium species, Coccidioides immitis, Histoplasma capsulatum, and others.

Portions of the 16S rRNA are conserved across many species of microorganisms. Amplifying the 16S rRNA using primers to these conserved regions allows isolation and sequencing of the variable regions of the molecules. These variable sequences are genus- or species-specific markers that allow identification of microorganisms. Pathogens that are difficult or impossible to culture in the laboratory have been identified using this technique. One example is Tropheryma whipplei, the cause of Whipple's disease.

·         CHOLERA:

HOLDING MEDIA:  VR Medium, Cary-Blair Medium, Autoclaved sea water

ENRICHMENT MEDIA: Allkaline Peptone water at pH 8.2, Monsur's Taurocholate tellurite peptone water at pH 9.2,

PLATING MEDIA: Alkaline Bile salt agar, Monsur's Gelatin taurocholate trypicase tellurite agar(GTTA) Medium, TCBS Medium.

V. cholerae O139 Bengal is in fact virtually identical to V. cholerae O1 El Tor except for two important differences: production of the novel O139 LPS and of an immunologically related O-antigen polysaccharide capsule. Encapsulation is not a feature of O1 strains and may explain the resistance of O139 strains to human serum in vitro as well as the occasional development of O139 bacteremia.