Figure 1.
Scanning electron micrographs (SEM) of the surface of mock-inoculated control trees at two different heights post-inoculation (PI).
(A) Surface of upper 15 cm stem segment, lacking any openings/lenticels, collected from the susceptible clone NC11505. (B) Surface of upper 15 cm stem segment, lacking any openings/lenticels, collected from the moderately resistant clone NM6. (C) Surface of lower 15 cm stem segment, with small openings/lenticels indicated by the arrow, of clone NC11505. (D) Surface of lower 15 cm stem segment, with small openings/lenticels indicated by an arrow, of clone NM6. Magnification and scale bars included on the bottom of each image.
Figure 2.
Scanning electron micrographs (SEM) of stems inoculated with a conidial suspension of Sphaerulina musiva at two different times (6 h and 12 h) post-inoculation (PI) depicting germination and penetration on the moderately resistant (NM6) and susceptible (NC11505) clones.
(A) SEM micrograph of stem surface of NC11505 depicting S. musiva conidium 6 h PI. (B) SEM micrograph of stem surface of NM6 and conidium of S. musiva 6 h PI. (C) SEM micrograph of stem surface of NC11505 with conidium and germ tube of S. musiva entering a lenticel 12 h PI. (D) SEM micrograph of stem surface of NM6 with conidium and germ tube of S. musiva penetrating a small opening 12 h PI. (E) Micrograph (C) at increased magnification depicting penetration of lenticel by a germ tube. (F) SEM micrograph of stem surface of resistant clone NM6 with germ tube of S. musiva penetrating a small opening 12 h PI. Tr = trichome, Sp = conidium, L = lenticel, GT = germ tube, Op = small opening. Magnification and scale bars included on the bottom of each image.
Table 1.
A comparison of the mean, range, and standard deviation of penetration rates on the moderately resistant clone NM6 (Populus maximowiczii×P. nigra) and susceptible clone NC11505 (P. maximowiczii×P. trichocarpa) inoculated with a spore suspension of Sphaerulina musiva.
Figure 3.
Bright field micrographs of transverse sections through 7-week-old mock-inoculated controls of the resistant (DN74) and susceptible (NC11505) clones depicting the gross anatomy of stem tissue.
(A) Stem anatomy of the mock-inoculated clone DN74. (B) Stem anatomy of mock inoculated clone NC11505. COX = Cortex, L = Lenticel, P = Periderm, PF = Phloem fiber. Scale bars = 200 µm.
Figure 4.
Transverse sections of susceptible clone NC11505 at 3 time points (3 weeks, 5 weeks, and 7 weeks) post-inoculation (PI) depicting anatomical responses to inoculation with a conidial suspension of Sphaerulina musiva.
(A) Bright field micrograph of necrotic stem lesion 3 weeks PI. Necrotic area (Nec) bounded by layer of impervious tissue (IT indicated with arrow). (B) Fluorescent micrograph of necrotic lesion 3 weeks PI. IT layer visible as light purple fluorescence (indicated with arrows). (C) Bright field micrograph of necrotic lesion 5 weeks PI. Nec is bounded by a layer of necrophylactic periderm (NP; indicated with arrow). (D) Fluorescent micrograph of necrotic lesion 5 weeks PI. Nec is bounded by IT layer and NP layer (indicated with arrows). (E) Bright field micrograph of necrotic stem 7 weeks PI. Entire cortex (COX) is necrotic and filled with collapsed cells. (F) Fluorescent micrograph of necrotic lesion 7 weeks PI. NP and IT are absent from the Nec. Blue auto-fluorescence viewed under ultraviolet light. Filter parameters: Excitation filter G 365, Beam Splitter FT 395, Emission filter BP 445/50. Green auto-fluorescence viewed under ultraviolet light. Filter parameters: Excitation filter BP 450–490, Beam Splitter FT 510, Emission filter BP 515–565. P = periderm. Pf = primary phloem fiber. X = Xylem. VC = Vascular cambium. Scale bars = 200 µm.
Figure 5.
Transverse sections of resistant clone DN74 at 3 time points (3 weeks, 5 weeks, and 7 weeks) post-inoculation (PI) depicting anatomical responses to inoculation with a conidial suspension of Sphaerulina musiva.
(A) Bright field micrograph of necrotic lenticel (L) 3 weeks PI. Necrophylactic periderm (NP; indicated by an arrow) is apparent immediately below the infected L at the margin between the cortex (COX) and Periderm (P). (B) Fluorescent micrograph of necrotic lenticel 3 weeks PI. NP layer is evident as thick square shaped cells fluorescing bright purple immediately below the lenticel within the COX (indicated by an arrow). (C) Bright field micrograph of necrotic lesion, bounded by NP, 5 weeks PI. (D) Fluorescent micrograph of two necrotic lesions 5 weeks PI. NP is evident as thick layer of purple fluorescing cells (indicated by arrows) immediately below the necrotic area (Nec). (E) Bright field micrograph of Nec 7 weeks PI. Nec is bounded by two successive layers of NP (indicated by arrows), with occluded xylem (X) cells (dark red to black cells in the X) adjacent to the Nec. (F) Fluorescent micrograph of necrotic lesion with two successive layers of NP appearing purple (indicated by arrows). Blue auto-fluorescence viewed under ultraviolet light. Filter parameters: Excitation filter G 365, Beam Splitter FT 395, Emission filter BP 445/50. Green auto-fluorescence viewed under ultraviolet light. Filter parameters: Excitation filter BP 450–490, Beam Splitter FT 510, Emission filter BP 515–565. P = periderm. Pf = Primary phloem fiber. VC = Vascular Cambium. Scale bars = 200 µm.
Figure 6.
Bright field micrographs of susceptible clone (NC11505), at three different time points (3 weeks, 5 weeks, and 7 weeks) post-inoculation (PI) with blue stained hyphae, indicated by arrows, visible in different tissues.
(A) Longitudinal section depicting hyphal growth in cortex at 3 weeks PI. (B) Transverse section through cortex depicting hyphal growth at 5 weeks PI. (C) Transverse section through xylem at 7 weeks PI depicting hyphal growth in xylem vessels (XV) and rays (XR). (D) Longitudinal section through xylem tissue depicting hyphal growth in vessels and rays. COX = Cortex, Pf = Primary phloem fiber, Pyc = Pycnidium, P = Periderm. Scale bars = 200 µm.