Salmonella Flipbook PDF

Salmonella Flipbook Prepared By: Matthew J. Forstner Laboratory Services MN Dept. of Agriculture This flipbook is intended as a general guide to aid analysts in the detection and identification of Salmonella growing on the plating media and screening tubes used in the BAM Chapter 5 Salmo...

Salmonella Flipbook Prepared By: Matthew J. Forstner Laboratory Services MN Dept. of Agriculture This flipbook is intended as a general guide to aid analysts in the detection and identification of Salmonella growing on the plating media and screening tubes used in the BAM Chapter 5 Salmonella method. For the purpose of this flipbook, “Typical” Salmonella refers to Salmonella enterica subsp. enterica isolates that produce H2S, are lysine decarboxylase positive, and do not ferment lactose or sucrose. When using this flipbook, please consider the following: The appearance of isolates encountered during sample analysis, whether typical or atypical, may vary from the images depicted. Atypical Salmonella can be rare. Some of the atypical morphologies depicted in this flipbook are so rare that they may not be encountered. Some serovars of Salmonella may be inhibited on bismuth sulfite agar. This flipbook does not address the phenotypic appearance of unique biovars of Salmonella, such as Salmonella ser. Typhi or Salmonella ser. Paratyphi A. Use caution if/when printing this flipbook, as the colors of the printed images may be distorted from those pictured on screen. Salmonella BAM Plate Morphology & Screening Tube Reactions  Typical Salmonella BAM Media HE XLD Green to blue—green H2S+ colonies. Opaque /yellow to red H2S+ colonies. TSI LIA TSI LIA Screening tubes  TSI  K/A H2S+  LIA  K/K H2S+ BS Brown, grey, or black colonies with darkening around colony.  H2S Negative Salmonella BAM Media HE XLD Green to blue - green H2S(-) colonies. Opaque/yellow to red H2S(-) colonies. TSI LIA TSI LIA Screening tubes  TSI  K/A H2S(-)  LIA  K/K H2S(-) BS BS Key features: H2S(-) colonies on HE and XLD Atypical morphology: Typical morphology: BS may be typical or atypical Green colonies, with little or no Brown, grey, or black colonies with H2S(-) TSI and LIA reactions darkening of the surrounding agar. darkening of the surrounding agar.  Lactose/Sucrose Positive Salmonella BAM Media HE XLD Orange or yellow colonies with or without H2S. Yellow colonies with or without H2S. TSI LIA TSI LIA Screening tubes  TSI  A/A H2S±  May or may not have slight red slant  LIA  K/K H2S± Key features: Orange colonies on HE H2S± Yellow colonies on XLD H2S± BS Brown, grey, or black colonies with darkening around colony. A/A reaction on TSI H2S±  Atypical BS Morphology Salmonella BAM Media HE XLD Green to blue—green H2S+ colonies. Opaque/yellow to red H2S+ colonies. Key Features: BS atypical HE and XLD both typical Screening tubes:  TSI  Isolates will be typical (K/A H2S+)  LIA  Isolates will be positive for H2S, BS Green colonies with little or no darkening of surrounding agar. but could be either K/K or K/A  Lysine Decarboxylase Negative Salmonella TSI LIA TSI LIA TSI LIA TSI LIA Lysine Decarboxylase Negative Lysine Decarboxylase Negative H2S Positive H2S Negative These rare isolates will appear typical on HE. These even more rare isolates will appear H2S On XLD, these isolates may appear typical, but negative on HE and XLD. On XLD, these isolates may also cause yellowing of surrounding agar. may cause yellowing of surrounding agar. On On BS, these isolates may be typical or atypical. BS, these isolates may be typical or atypical.  Hektoen Enteric Agar Salmonella Morphologies H2S Negative H2S Positive Typical H2S Negative Lactose/Sucrose Positive  Xylose Lysine Desoxycholate Agar Salmonella Morphologies Xylose Positive H2S Negative Xylose Negative H2S Positive Typical H2S Negative Lactose/Sucrose Positive Note: Salmonella are typically xylose positive, which results in colonies with a dull yellow color after 22-26 hours of incubation, instead of the “pink colonies” described by the BAM. However, atypical xylose negative Salmonella will produce pink – red colonies on XLD after 22-26 hours of incubation. The yellow color caused by xylose fermentation is less intense than the yellow color expressed from atypical lactose and/or sucrose fermenting isolates. Note: With prolonged incubation, or even letting the plates sit out on the bench, the yellow color caused by xylose fermentation, changes to a pink to red color.  Bismuth Sulfite Salmonella Morphologies Typical Morphology Atypical Morphology  Bismuth Sulfite Salmonella Morphologies at 24 & 48 Hours Example 1 at 24 Hours Example 2 at 24 Hours Example 3 at 24 Hours Example 1 at 48 Hours Example 2 at 48 Hours Example 3 at 48 Hours Note: Bismuth sulfite agar must be analyzed after 24 and 48 hours of incubation. In some cases, Salmonella may form typical colonies after 24 hours of incubation, as shown in Example 1. However, it is not uncommon for Salmonella to only form typical colonies after 48 hours of incubation, as shown in Example 2. In some cases, Salmonella may not form typical colonies at all, as shown in Example 3.  Evaluation of Suspicious Salmonella Colonies Examine HE, XLD, and BS plates for presence of colonies that may be Salmonella. BS plates should be examined after 24 ± 2 hours and returned to incubator to be examined again after 46 to 50 hours total incubation. XLD Typical Salmonella appear as opaque/yellow, pink, or red colonies with black centers. Look for colonies with a slightly rough or dimpled H2S center. Atypical Salmonella  H2S negative Salmonella appear as opaque/yellow, pink, or red colonies without black centers.  Lactose or sucrose fermenting Salmonella appear as yellow colonies with or without black centers. HE Typical Salmonella appear as blue-green to blue colonies with black centers. Many Salmonella cultures have large glossy black centers or may appear as almost completely black colonies. Atypical Salmonella  H2S negative Salmonella appear as blue-green to blue colonies without black centers.  Lactose or sucrose fermenting Salmonella appear as yellow to orange colonies with or without black centers. BS Typical Morphology  Salmonella appear as brown, gray, or black colonies that may have a metallic sheen. Surrounding media is usually brown at first, turning black with increased incubation time. Atypical Morphology  Salmonella strains produce green colonies with little or no darkening of surrounding media.  Evaluation of Suspicious Salmonella Isolates Salmonella reactions on TSI and LIA tubes. TSI Typical Salmonella will produce alkaline (red) slants and acid (yellow) butts with hydrogen sulfide (black). K/A + Atypical Salmonella  H2S negative Salmonella will produce alkaline (red) slants and acid (yellow) butts without hydrogen sulfide (black). K/A (-)  Lactose or sucrose fermenting Salmonella will produce both acid (yellow) slants and butts or have a slight red slant (SRS) with the balance of the tube yellow, with or without hydrogen sulfide (black). A/A ± with or without SRS LIA Typical Salmonella will produce both alkaline (purple) slants and butts with hydrogen sulfide (black). K/K + Atypical Salmonella  H2S negative Salmonella will produce both alkaline (purple) slants and butts without hydrogen sulfide (black). K/K (-)  Lysine decarboxylase negative Salmonella will produce alkaline (purple) slants and acid (yellow) butts, with or without hydrogen sulfide (black). K/A ±  TSI & LIA Rule-Out Reactions Isolates giving any of the following reactions may be considered negative for Salmonella: 1. Isolates producing a “no change”, or alkaline slant and butt (K/K) in both TSI (red) and LIA (purple). 2. Isolates with the “three A’s you’re out” reaction. TSI = A/A LIA= acid (A) butt 3. Isolates producing a burgundy or brick red slant in LIA. R/A