【文章內(nèi)容簡介】 differentiate them). ? Growth on selectivedifferential media, such as SalmonellaShigella (SS) medium, eosinmethyleneblue (EMB) and MacConkey agar ? The select effect of the media in suppressing unwanted grampositive anisms is exerted by bile salts or bacteriostatic dyes in the agar. ? The differential ability of these media is based on lactose fermentation: normal flora positive ( colored colonies) and pathogens negative (colorlesscolonies). Separation of bacteria : plate streaking. Bacteriologic plate streaking. ? Colony: the visible growth of bacteria on solid growth media. Ideally, the colony is the progeny of one, or at most, a few bacteria. ?A colony will usually contain millions of bacterial cells. ?Colony morphology can sometimes be useful in bacterial identification. ?Colonies are described as to such properties as size, shape, texture, elevation, pigmentation, effect on growth medium. Colony Growth on blood agar to test for hemolytic properties α hemolytic: inplete lysis of red blood cells, resulting in a greenish halo around the colony β hemolytic: plete lysis of red blood cells, resulting in a clear halo around the colony ?hemolytic: nonhemolytic 1. Cultural characteristics: ? unique nutritional requirements, ? pigment production, ? hemolytic properties Identification of Bacteria ? a (alpha) ? partial hemolysis ? greenish color (. S. pneumoniae) ? b (beta) ? plete clearing ? Group A and B ? ? (gamma) ? no lysis ? Enterococcus (groupD) hemolysis reaction sheep blood agar MacConkey’s Agar Contains lactose and a pH indicator, E. coli ferment lactose to produce acid, which turns the pH dye red. So, E. coli colonies appear red. 2. Biochemical characterization Identification of Bacteria ? the ability to attack various substrates ? or to produce particular metabolic products 2. Biochemical characterization Triple Sugar Iron Agar Identification of Bacteria A simple approach to rapid diagnosis (as an example of antigen detection) is used in many doctor39。s offices for the group A streptococcus. The patient39。s throat is swabbed and streptococcal antigen extracted directly from the swab (without prior bacteriological culture). The bacterial antigen is detected by aggregation (agglutination) of antibody coated latex beads. Serologic identification of an antibody response (in patient39。s serum) to the infecting agent can only be successful several weeks after an infection has occurred. Identification use of antibodies of known specificity to detect antigens present on whole bacteria or free in bacterial extracts Identification of Bacteria Serological methods – used to detect both antigen and antibody in specimens ? The fastest and most specific way ? Immunofluorescence – microscopy, FACS ? Enzymeimmunoassay – ELISA, Western blot ? Radioimmunoassay – quantitate antigenantibody plex FACS (fluorescence activated cell sorter) 4. Genomic Methods ? 16S DNA sequencing ? Labeled probes specific for the 16S rRNA of a species are added, and the amount of label on the doublestranded hybrid is measured. ? This technique is widely used for the rapid identification of many anisms. ? Fluorescence in situ hybridization (FISH) ? Polymerase Chain Reaction (PCR) ? Nucleic Acid Sequence Analysis ? Checker board DNA hybridization (DNA microarray) Identification of Bacteria ELISA (EnzymeLinked Immunosorbent Assay) E. coli 16S RNA ? The 16S rRNA of each species of bacteria has stable (conserved) portions of the sequence. ? Many copies are present in each anism. FISH PCR (polymerase chain reaction) Bacteria mRNA cDNA DNA microarrays 37oC – cDNA labeled w/ fluorescein tag 25oC – cDNA labeled w/ rhodamine tag DNA array ~6000 genes “Transcriptome” Summary ? Direct microscopy ? Fast ? Give some hints on the type of bacteria ? Low sensitivity ? Culture ? High sensitivity ? Can make definitive ID ? Slow ? Only works on culturable bacteria ? Serological assay ? Fast ? High specificity ? Can detect both antigen and antibody ? Easy (can be used at chairside or bedside) ? Low sensitivity ? Genomic based assay ? Fast ? High sensitivity and specificity ? Works on both culturable and nonculturable bacteria ? Especially useful for detecting slow growing bacteria such as . ? Requires technical expertise ? False positive or negative Prevention of Bacterial Infection Artificial active immunity Vaccines are antigens prepared from pathogens that can raise a protective immune response, yet do not cause illness. These prepared antigens will stimulate both B cells and T cells and help to create memory cells that can later mount a vigorous immune response to an encounter with the real pathogen. ? Toxoids: a modified form of the toxin that preserves its antigenicity but has lost its toxicity. This has been spectacularly successful with tetanus and diphtheria. ? Inactivated vaccines: ? Attenuated live vaccines : ? Special vaccines: polysaccharide vaccine, subunit vaccine, ( conjugate vaccine, bioengineered vaccine, chemical vaccine, synthetic vaccine ), nucleic acid vaccine, idiotype vaccine, autovaccine, etc. Artificial active immunity Artificial passive immunity ? Antitoxin: . Tetanus antitoxin and diphtheria antitoxin. It is raised in the horse. It is most important to give an intented recipient of equine serum a prior test dose to exclude hypersensitivity subjects who may have been sensitized by a previous dose of equine serum. ? Pooled immunoglobulin: It contains the normal repertoire of antibodies for an adult, and can protect against hepatitis A, and measles. ? Specific immunoglobulin: Preparations of specific immunoglobulin are available for passive immunization against t