Figure 1.
Clinical and Pathological Images of the Patient's Lymph Nodes
(A) MRI of the chest showing adenopathy in the precarinal, subcarinal, and hilar regions. Central areas of diminished enhancement suggest necrosis (arrow).
(B) Removed cervical lymph node showing necrotizing granulomatous lymphadenitis with abscess formation (pyogranuloma).
(C) Warthin-Starry stain of the cervical lymph node (magnification 600×) showing coccobacillary organisms.
(D) Higher magnification H&E of a necrotizing granuloma. There is an area of neutrophils and cellular debris on the right, bounded by a poorly defined layer of palisaded epithelioid histiocytes. Outside this layer is a mix of lymphocytes and histiocytes.
Figure 2.
Novel Bacterium Groups in the Family Acetobacteraceae
(A) Phylogenetic tree derived from the 16S rDNA sequence of the patient isolate, two Gluconacetobacter spp, two Acetobacter spp., and other gram-negative rods associated with CGD (S. marcescens, B. cepacia) and not associated with CGD (Escherichia coli). This unrooted radial tree shows the patient isolate emerging as an independent branch that clustered together with members of the family Acetobacteraceae. The scale bar represents the number of substitutions per site.
(B) Percentage of similarity of 16S rDNA sequences of the patient isolate, two Gluconacetobacter spp., two Acetobacter spp., and the gram-negative rods shown in (A).
Figure 3.
Immune Response to G. bethesdensis
(A) An immunoblot showing the patient's plasma incubated with whole bacterial protein extracts of this new pathogen and other organisms as indicated (2 μg total protein per lane). Patient plasma was used at a 1:1000 dilution.
(B) Patient's plasma from 1999 (a) and 2003 (b) incubated with whole bacterial protein extracts.
Figure 4.
The Amino Acid Sequence of the Methanol Dehydrogenase
Methanol dehydrogenase (69 kDa; EC 1.1.99.8) was identified as one of the peptides recognized by the patient plasma. Amino acids in red represent sequences that were matched during peptide mass fingerprinting.
Figure 5.
Immune Electron Microscopy of the New Bacterium
Bacteria were adsorbed to Parlodion-coated grids and then probed with either normal plasma (A) or plasma from the CGD patient (B), labeled with anti-human IgG/colloidal gold, and negatively stained with uranyl acetate.
Figure 6.
Histopathology of Mouse Lymph Nodes
(A) Normal lymph node architecture of a gp91phox−/− uninjected control mouse (magnification 5×).
(B) Normal lymph node architecture of a gp91phox−/− mouse after 107 i.p. inoculation of Gluconobacter oxydans (magnification 5×).
(C) Moderate lymphoid hyperplasia and presence of atypical macrophages in the lymph node of a gp91phox−/− mouse after 106 intraperitoneal inoculation of the novel organism (magnification 5×).
(D) View of the epithelioid macrophages in the lymph node at 40×.
(E) Lymphadenitis in the cervical lymph node of a gp91phox−/− mouse after 107 i.p. inoculation of the novel organism (magnification 5×).
(F) View of the pyogranulomatous reaction in the lymph node at 20×.
(A,B,E,F) are from mice that were sacrificed at nine days post-inoculation. (C and D) are from a mouse that was sacrificed at 76 days post-inoculation.
Figure 7.
Graph Showing Organism Recovery from Spleen Homogenates at Nine Days Post-Inoculation
The — mark shows the mean value. Mean CFU/spleen in the wild-type mice was 11,550, compared with 41,720 in the knockout animals (p < .001; two-tailed t-test).