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Figure 1: Evidence Shirt Illustrating the Presence of Nitrates. This evidence shirt illustrates the presence of nitrites with a positive test for gun shot residue. The chemistry involved is the same used in the microbiology laboratory to detect nitrate (NO3) reduction to nitrite (NO2).  (J. Scott Doyle, Jefferson Regional Forensic Lab, Louisville, KY)

Figure 2: Positive Greiss Test Illustrating the Presence of Nitrites. This positive Greiss test illustrates the presence of nitrites. The chemistry involved is the same used in the microbiology laboratory to detect nitrate (NO3) reduction to nitrite (NO2).  (J. Scott Doyle, Jefferson Regional Forensic Lab, Louisville, KY)

Figure 3: Uninoculated Tube. Uninoculated tube illustrating the proper filling of nitrate and nitrite broth. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 4: Nitrate Reagents A and B. Nitrate reagents A and B. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 5: Zinc dust. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 6: "Dirtying" a stick with zinc dust. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 7: Addition of Zinc Dust to a Nitrate Test. Addition of zinc dust to a nitrate test to force the reduction of the substrate. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 8: Addition of Zinc Dust to a Nitrate Test (Labeled view). Addition of zinc dust to a nitrate test to force the reduction of the substrate. (Labeled view) (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 9: Negative Nitrate Reaction After the Addition of Reagents A and B. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 10: Negative Nitrate Reaction After the Addition of Reagents A and B and Zinc Dust. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 11: Negative Nitrite Reaction After the Addition of Reagents A and B. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 12: Pseudomonas aeruginosa growing in nitrate and nitrite broths before the addition of reagents, illustrating gas production. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 13: Pseudomonas aeruginosa growing in nitrate (tube labeled NO3) and nitrite (tube labeled NO2) broths before the addition of reagents, illustrating gas production. (Labeled view) (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 14: Pseudomonas aeruginosa growing in nitrate and nitrite broths after the addition of reagents A and B. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 15: Pseudomonas aeruginosa growing in nitrate (tube labeled NO3) and nitrite (tube labeled NO2) broths after the addition of reagents A and B. (Labeled view) (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 16: Pseudomonas aeruginosa growing in nitrate and nitrite broths after the addition of reagents A and B and zinc dust. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 17: Pseudomonas aeruginosa growing in nitrate (tube labeled NO3) and nitrite (tube labled NO2) broths after the addition of reagents A and B and zinc dust. (Labeled view) (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 18: Moraxella catarrhalis growing in nitrate and nitrite broths before the addition of reagents. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 19: Moraxella catarrhalis growing in nitrate (tube labeled NO3) and nitrite (tube labeled NO2) broths before the addition of reagents. (Labeled view)  (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 20: Moraxella catarrhalis growing in nitrate and nitrite broths 24 hours after the addition of reagents A and B. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 21: Moraxella catarrhalis growing in nitrate (tube labled NO3) and nitrite (tube labeled NO2) broths 24 hours after the addition of reagents A and B. (Labeled view) (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 22: Moraxella catarrhalis growing in nitrate and nitrite broths 24 hours after the addition of reagents A and B and zinc dust, showing partial reduction of nitrate. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 23: Moraxella catarrhalis growing in nitrate (tube labeled NO3) and nitrite (tube labeled NO2) broths 24 hours after the addition of reagents A and B and zinc dust, showing partial reduction of nitrate. (Labeled view) (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 24: Moraxella catarrhalis growing in nitrate and nitrite broths 48 hours after the addition of reagents A and B and zinc dust, showing complete reduction of nitrate. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 25: Moraxella catarrhalis growing in nitrate (tube labeled NO3) and nitrite (tube labeled NO2) broths 48 hours after the addition of reagents A and B and zinc dust, showing complete reduction of nitrate. (Labeled view) (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 26: Escherichia coli growing in nitrate and nitrite broths before the addition of reagents. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 27: Escherichia coli growing in nitrate (tube labeled NO3) and nitrite (tube labeled NO2) broths before the addition of reagents. (Labeled view) (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 28: Escherichia coli growing in nitrate and nitrite broths after the addition of reagents A and B. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 29: Escherichia coli growing in nitrate (tube labeled NO3) and nitrite (tube labeled NO2) broths after the addition of reagents A and B. (Labeled view) (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 30: Neisseria lactamica growing in nitrate and nitrite broths before the addition of reagents. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 31: Neisseria lactamica growing in nitrate (tube labeled NO3) and nitrite (tube labeled NO2) broths before the addition of reagents. (Labeled view) (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 32: Neisseria lactamica growing in nitrate and nitrite broths after the addition of reagents A and B. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 33: Neisseria lactamica growing in nitrate (tube labeled NO3) and nitrite (tube labeled NO2) after the addition of reagents A and B. (Labeled view) (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 34: Neisseria lactamica growing in nitrate and nitrite broths after the addition of reagents A and B and zinc dust. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 35: Neisseria lactamica growing in nitrate (tube labeled NO3) and nitrite (tube labeled NO2) broths after the addition of reagents A and B and zinc dust. (Labeled view) (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 36: Composite slide illustrating the proper filling of nitrate or nitrite broth with an extended legend. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 37: Composite slide illustrating the reagents used to visualize nitrate and nitrite reduction with an extended legend. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 38: Composite slide illustrating the addition of zinc dust to nitrate and nitrite reduction reactions with an extended legend. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 39: Composite slide illustrating a negative nitrate reaction with an extended legend. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 40: Composite slide illustrating a negative nitrite reaction with an extended legend. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 41: Composite slide illustrating nitrate and nitrite reactions for Pseudomonas aeruginosa with an extended legend. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 42: Composite slide illustrating nitrate and nitrite reactions for Moraxella catarrhalis with an extended legend. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 43: Composite slide illustrating nitrate and nitrite reactions for Moraxella catarrhalis with an extended legend. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 44: Composite slide illustrating nitrate and nitrite reactions for Escherichia coli with an extended legend. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 45: Composite slide illustrating nitrate and nitrite reactions for Neisseria lactamica with an extended legend. (Rebecca Buxton, University of Utah, Salt Lake City, UT)

Figure 46: Alcaligenes faecalis grown in nitrate broth. Nitrate reagents A and B were added to the broth after 48 hours incubation; no color change was observed. Zinc granules were added, producing a red color change in the broth.  The zinc particles forced the reduction of nitrate to nitrite. (Diane Hartman, Baylor University, Waco, TX)

Figure 47: Escherichia coli grown in nitrate broth.  Nitrate reagents A and B were added to the broth after 48 hours incubation. The red color indicates reduction of nitrate to nitrite. (Diane Hartman, Baylor University, Waco, TX)

Figure 48: Pseudomonas aeruginosa grown in nitrate broth. Nitrate reagents A and B were added to the broth after 48 hours incubation; no color change was observed. Zinc granules were added, and the broth remained golden. Nitrogen bubbles rose to the surface of the broth, indicating denitrification. (Diane Hartman, Baylor University, Waco, TX)

Figure 49: Nitrate broths after the addition of reagents A (sulfanilic acid in acetic acid) and B (nn-dimethyl-1-naphthylamine in acetic acid).  All cultures were incubated at 37°C for 48 hours before the addition of each reagent (10 drops each).  A)  Alcaligenes faecalis showed no color change indicating that zinc must be added to complete analysis.  B) Escherichia coli  turned red after the addition of reagents A and B indicating that nitrate has been reduced to nitrite.  Zinc does not need to be added to complete analysis.  C) Pseudomonas aeruginosa showed no color change indicating that zinc must be added to complete analysis.  (Tasha Sturm, Cabrillo College, Aptos, CA)

Figure 50: Nitrate broths after the addition of reagents A (sulfanilic acid in acetic acid) and B (nn-dimethyl-1-naphthylamine in acetic acid) and zinc dust.  All cultures were incubated at 37°C for 48 hours before the addition of reagents A and B (10 drops each) and zinc dust. A) Alcaligenes faecalis turned red, indicating that nitrate is still present and has not been reduced (nitrate reacts with zinc). B) Escherichia coli turned red after the addition of reagents A and B, indicating that nitrate has been reduced to nitrite. Therefore, zinc was subsequently not added.  C) Pseudomonas aeruginosa showed no color change indicating that nitrate has been completely reduced beyond nitrite to ammonia. (Tasha Sturm, Cabrillo College, Aptos, CA)

Figure 51: Nitrate Test. The nitrate test on the left shows a positive reaction after addition of reagents A (sulfanilic acid in acetic acid) and B (nn-dimethyl-1-naphthylamine in acetic acid).  The nitrate test on the right shows a negative reaction. (Ken Van Horn, Focus Diagnostics, Inc., Cypress, CA)

Figure 52: Nitrate Reduction Test Results Following Growth in Nitrate Broth. Nitrate reduction test results following growth of Escherichia coli, Alcaligenes faecalis, and Pseudomonas aeruginosa in nitrate broth. Escherichia coli reduces nitrate to nitrite, as evidenced by the red color after addition of nitrate reagents A (sulfanilic acid) and B (dimethyl-α-naphthylamine) to the culture. Alcaligenes faecalis is negative for nitrate reduction as the culture turns red only after addition of reagents A, B, and zinc powder. Pseudomonas aeruginosa denitrifies nitrate to nitrogen gas. Note the gas bubbles captured by the Durham tube (arrow) and visible at the top of the broth. An uninoculated nitrate broth is shown for comparison. (Andrew Herbig, Washburn University, Topeka, KS) 

Figure 53: Nitrate Reduction Test Results Following Growth in Nitrate Broth (Labeled view) (Andrew Herbig, Washburn University, Topeka, KS)