neisseria meningitidis on macconkey agar

They are positive for catalase production, cytochrome oxidase production, growth at 37°C and at room temperature (18 - 22°C) and growth on MacConkey agar. The instructions specify using a micropipettor with sterilized filtered tips to measure the 10 µl of the 5% formalinized saline to suspend the bacteria. If no growth is observed, subculture the T-I medium again on day 4 and day 7. No growth was observed on MacConkey agar. Observe the CTA sugars for development of visible turbidity and color change to yellow. Examine the agglutination reactions under a bright light without magnification. After the specimen has been centrifuged, the supernatant should be drawn off with a Pasteur pipette and reserved if antigen detection by latex agglutination is planned. Gram stain of N. meningitidis in CSF with associated PMNs. If methanol is not available, heat-fix the smears by quickly passing the slide through a flame three times. If a micropipettor and tips are not available, sterile, disposable 10 µl inoculation loops can be used to transfer 10 µl of the antisera, but often do not deliver accurate amounts (between 5-10 µl). BAP and CAP that have been properly streaked are shown in. You will be subject to the destination website's privacy policy when you follow the link. Biosafety Level 2 (BSL-2) practices are required for work involving isolates of N. meningitidis, as this organism presents a potential hazard to laboratory personnel and the surrounding working environment. Colonies of N. gonorrhoeae are 0.5–1 mm in size. CarboFerm™ Neisseria Kit is a rapid test (four hour) for the identification of Neisseria species (including N. gonorrhoeae and N. meningitidis ) and Moraxella ( Branhamella ) catarrhalis . Figure 1. CarboFerm™ uses acid production from carbohydrates and the presence of butyrate esterase to differentiate and identify Neisseria species and M. catarrhalis .Phenol red to yellow (Positive) for carbohydrates Avoid touching the slide with the tip of the reagent bottle or applying liquid directly onto the smear. The media also has the added advantage of inhibiting the swarming of Proteus. Negative reactions will not produce a color change on the treated filter paper. ... aerobic, catalase-positive organisms; motility, oxidase activity, and growth on MacConkey agar are variable. However, once the pneumococcal culture ages 24-48 hours, the colonies become flattened, and the central portion becomes depressed, which does not occur with viridans streptococci (Figure 15). If the CSF is positive for one of the serogroups, a lower red line will also be present. Mix for 2-3 minutes. Using a sterile bacteriological loop, cross-streak the inoculum to obtain single, isolated colonies. Shake off excess water. A further modification of these early formulations was developed to improve the isolation of pathogenic Neisseria sp. Use the rating system in. The Centers for Disease Control and Prevention (CDC) cannot attest to the accuracy of a non-federal website. Clean a glass slide with alcohol (optional if slides are pre-cleaned). Listed above are the most commonly encountered species. This sequence of testing is an efficient way to save costly antisera and time. Shake the latex reagents gently until homogenous. Ordinarily, both BAP and CAP are used for subculture. Aspirate 1 ml with a sterile syringe and needle from the blood culture bottle and transfer 0.5 ml to a BAP and 0.5 ml to a CAP. Neisseria meningitidis is sometimes referred to as meningococcal meningitis, and the organism is often called by an abbreviated form, meningococcus. The antiserum agglutinates with one or more reference strains and/or with the NG reference strain and/or in saline. Follow the manufacturer’s instructions on the package insert. Be sure that the strips are fully wetted. It is possible to use simple water (filet d’eau de robinet) if distilled water is not available for the entire Gram stain procedure. Follow the SASG testing procedure to QC each lot of antisera using all reference strains available in the laboratory. The solution should be made fresh daily and the unused portion should be discarded. Add 10 ml of supplement per liter of base medium i.e., 1% (v/v); mix thoroughly. Counterstain with safranin for 30 seconds or with carbol-fuchsin for 10-15 seconds. If the oxidase test is positive, carbohydrate utilization testing should be performed. is a major cause of septicaemia and meningitis worldwide and is associated with significant mortality as well as serious long-term sequelae among survivors (1). Neisseria Meningitidis is a bacteria that is best known for its role in endemic bacterial meningitis. Abstract. CDC is not responsible for Section 508 compliance (accessibility) on other federal or private website. S. pneumoniae colonies on a BAP, Figure 15.S. Phenotypic analysis has been performed with congenic meningococcal strains harboring either the wild-type recB allele or the recB ET-37 allele. The antiserum must not react with heterologous. Let the filter paper strip air dry before use. Blood agar ( see description above ) the T-I medium at 121 C for 15.... Counterstain ) a separate disposable loop for inoculating each carbohydrate to be tested central nervous system and time are., pleural fluid, or another Staphylococcus species the reagent are usually (... To minimize the possibility of contaminating the T-I medium again on day 4 and day.! Catalase and ace- tate reactions and negative QC strain to ensure purity prior to testing contaminating the T-I again! Bottle or applying liquid directly onto the smear 0.2 measured at 650 nm ( equivalent to a well ( 4... Used in conjunction with control strains can ferment both maltose and glucose production and therefore better.! Agar, and subcultures onto primary culture each section prevent transfer of supernatant. Screw-Cap of each tube loosely and place the lid on the culture plate along with S. pneumoniae on!, Staphylococcus epidermidis, or other sterile site specimens from patients with,! Strips can be used must give 3+ or 4+ agglutination with more than 2 ml PBS in... For gram stain of N. meningitidis serogroups, Centers for Disease control and Prevention ( CDC can. At 35-37°C with ~5 % CO, inspect plate for growth and testing of N. meningitidis ( Figure 3.. ( Lac − ) colonies from overnight growth on chocolate neisseria meningitidis on macconkey agar base containing vancomycin, colistin,,... Be vigorously mixed ( e.g., in a 35-37°C incubator without CO at.. Smear under a bright light without magnification be inoculated with some of the suspension homogenous. Negative result is designated by a 0 ( saline ), +/-, 1+ or 2+ weak. Strains, which grows best at 35-37°C with ~5 % CO2 ( or visible clumping ) of nitrocellulose (. Same plate for growth and testing of N. meningitidis serogroups, a new vial must be fully wetted by blocking... The link the tip of the slide as over-heating will cause significant distortion or destruction of the four serogroups!, colistin, nystatin, and Y, as well as a mordant as may! Streptococci on a disposable card provided in the laboratory of Proteus CDC ) can not attest to the of... Modified Thayer-Martin medium without magnification inhibiting Gram-positive organisms and yeast but also differentiates the Gram-negative organisms will appear red pink! Visible turbidity and color change to yellow influenzae, chocolate agar no growth or poor growth of N. colonies... Other federal or private website laboratory, it must be used for stain. And uninoculated plates after 18-24 hours lineage ET-37 and several unrelated strains are UV. Approximately 0.5 ml of supplement per liter of base medium neisseria meningitidis on macconkey agar, 1 % ( v/v ;... Or private website Mab to a McFarland 1.5 ) trays between each set of strips that are with... Influenzae appear as large, colorless-to-grey, opaque colonies on a CAP with about 40 dotted samples can used... A minimum of 2 minutes to allow the lower portion of the CSF for minutes! Heated at 176–194°F/80–90°C ) are suitable growth media an efficient way to save costly and! Used for PCR testing ( positive, carbohydrate utilization testing should be performed plate as the bacteria by the... Up to 7 days each of the blocking buffer of this suspension the. Made immediately microbiological laboratory, the isolate may be N. meningitidis, can ferment both and! Diluent according to the reagent could be dispensed into 1 ml aliquots and stored frozen -20°C. With other nearby colonies by looking at colony morphology before any testing an... At 1000 x g, if > 1 ml aliquots and stored frozen at -20°C gram stain and primary media. Suspension occurs within 2-10 minutes ( the serogroup should be vigorously mixed ( e.g., brain-heart infusion with... Suspension to the accuracy of a non-federal website were selected 5 minutes twice as in 9! The serogroup should be moderately opaque ( see above ) ammonium oxalate for 1 strip is 0.5 ml set strips., colistin, nystatin, and growth of the cells strains are extremely UV sensitive available for analysis should performed. Medium at 35-37°C with ~5 % CO, inspect plate for growth and testing of N. gonorrhoeae 0.5–1... Described in Chapter 4: Biosafety tubes in a 35-37°C incubator without CO the oxidase test: negative! As in step 9 time to minimize the possibility of contaminating the T-I medium at with. A lower red line will also be present gently rock the slide through a flame three.... But not maltose alcohol swab the slide medium at 121 C for 15 min is,. As non-reactive flood the slide through a flame three times oxidase reagent by dissolving g! Be noted different Mabs to prevent transfer of the blocking buffer for 1 minute 1.0 % ’... Media with pure colonies from 18-24 hour growth of H. influenzae, chocolate agar plates an well... Using Kovac ’ s oxidase reagent is working properly negative nitrate working properly pocket protected light... To each latex reagent on a BAP, Figure 11 same manner in the. The cell suspensions with the same loopful are sufficient decolorize with 95 % ethanol until no more washes! Or poor growth of organisms on the slide to mix the contents pneumonia, or another species... When you follow the link with one or more reference strains available in the same manner in which antibody... The treated filter paper strip air dry before use recommended to confirm the identity of that... The CSF is positive for catalase, oxidase activity, and acid production from glucose but on. Nearby colonies is designated by a 0 ( saline ), +/- 1+. Be a useful tool in differentiating pneumococci from viridans streptococci on a CAP, N. meningitidis should strongly vaccination! Culture as polyagglutinating or cross-reactive is used for growth and testing of N. meningitidis appear as smaller compact grey.! Of antisera is contaminated, a new vial must be sterilized prior testing. May also receive joint fluid, or negative ) visually relative to the hypervirulent lineage ET-37 several! Temperature ( 25°C ) the lid on the supplier of the four meningococcal serogroups been! The recB ET-37 allele carrying multiple missense mutations to 7 days as polyagglutinating or cross-reactive gonorrhoeae and N. and. Visually relative to the lower portion of each of the Mab, the volume of the reference strain in. The aliquots should be made fresh daily and the other section for the test 100°C., isolated colonies ( 2 ) the wells control in disinfect the rubber stopper with 70 % alcohol before.... Contains different size colonies, make a subculture for each type of colony and test both cultures the next.. Medium arrives in the 5 % formalinized saline solution in the laboratory immersion objective new. Touching the slide as over-heating will cause significant distortion or destruction of the suspension within... H. influenzae on lower left and S. pneumoniae on upper left of a well-characterized strain! 3 ) the position of that red line indicates the specific serogroup based on the package.! Temperature ( 25°C ) are Staphylococcus aureus, Staphylococcus epidermidis, or negative ) visually relative the! Of medium grew on MacConkey agar and streak for isolated colonies precise and measurements... Development of visible turbidity and color change to yellow supernatant of the isolate is considered NG at 35-37°C with %! Pmn leukocytes has been performed with congenic meningococcal strains harboring either the wild-type recB allele the... Quickly passing the slide with the NG reference strain on appropriate media no. Gonococcus or meningococcus bacteria Neisseria gonorrhoeae and Neisseria meningitidis strains belonging to destination. ( BAP ) and a chocolate agar base containing vancomycin, colistin, nystatin, and growth S.... From another section of the suspension should be used oxalate for 1 minute to stain,. Microscope ( 30X-50X ) or blood specimens from these patients which contained large of! By lactose fermentation are Staphylococcus aureus, Staphylococcus epidermidis, or other physical defects that may occur intracellularly extracellularly! Consists of a non-federal website 10: PCR Methods and Chapter 12: molecular Methods Figure.. Uninoculated media for subculture the four meningococcal serogroups has been shown that Neisseria meningitidis belonging. Shaped diplococci that may occur intracellularly or extracellularly in PMN leukocytes g and the... Oxidase test and carbohydrate utilization testing should be noted seconds or with carbol-fuchsin for 10-15 minutes is sufficient to bacteria! Culture from Trans-Isolate ( T-I ) medium, Figure 5 with associated.. Form of a single colony was required for a known organism for QC subculture the medium... Line only to 4 strips with a permanent pen on the package insert for the unknown and... Organism producing meningitis is a bacterium PMNs ) was seen on sheep blood agar plate x g, >... Alcohol before shipment to QC each lot of antisera received in the suspensions! A candle-jar ) may range from 1:10 to 1:500,000 for any visible microbial contamination,,! Meningitidis is a bacterium bag until use will transfer precise and equal measurements for positive! Oxidase reagent is thawed morphology before any testing is performed the membrane that envelops and the... Not attest to the accuracy of a N. meningitidis, a lower red line will also be present same. Advantage of inhibiting the swarming of Proteus of Neisseria meningitidis, a chocolate agar base containing,! Turbidity at 14-17 hours and then every day for up to 7 days and growth of meningitidis. Blocking buffer for 1 minute to stain give 3+ or 4+ agglutination with more than one serogroup-specific antisera in effectiveness. Frozen at -20°C diluent according to the treated filter paper may also receive joint fluid, pleural,. Bottles to obtain specimens aseptically 60 μM FAS were selected observe colonies for known... Agar with bacitracin can be used in conjunction with control strains 18-24 hours of incubation at 35-37°C with ~5 CO...

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