Use of infrared microspectroscopy to determine leaf biochemical composition of cassava in response to Bacillus subtilis CaSUT007

† First and second authors are equally contributed to this work.

Figures & data

Figure 1. Effects of PGPR Bacillus subtilis CaSUT007 on the growth and development of cassava seedling under greenhouse conditions, as measured at 14 days after inoculation. (a) cassava seedling under greenhouse conditions in sterile soil, (b) percent of seedling germination, (c) dry weight, and (d) number of lateral root.

Note: The data are the average of four replications (three plants per replication) for each treatment. Error bars represent the standard deviation. For each growth parameter, different letters indicate significant differenced (P < .05) among treatments.

Figure 2. Functional group area maps of a part of a transverse leaf section from cassava achieved using FTIR microspectroscopy point to point mapping with aperture setting at 15 × 15 µm square aperture with 7.5 µm steps. (a) A micrograph of the untreated (control) transverse section. (b) A micrograph of a transverse cassava leaf section from CaSUT007 treated plants. (c) Integrated functional groups maps obtained under the spectral region between 1700–1580 cm^–1 for untreated (control). (d) Cassava leaf treated with CaSUT007. Dimensions of the area studied of 120 × 100 µm.

E, Epidermis; M, Mesophyll; VB, Vascular bundle.

Figure 3. FTIR mapping of a cassava leaf (control). (a) 2D HCA map performed using five clusters on the range of 1800–900 cm^–1. Different clusters are encoded by different colors. (b) Representative original average spectra of the spectra cluster. (c) Representative second derivative average spectra of the spectra cluster. Mesophyll (cluster 1), vascular bundle (cluster 2 and 3), and epidermis (cluster 4 and 5).

Figure 4. FTIR mapping of a cassava leaf section from CaSUT007 treated plants. (a) 2D HCA map performed using five clusters on the range of 1800–900 cm^–1. Different clusters are encoded by different colors. (b) Representative original average spectra of the spectra cluster. (c) Representative second derivative average spectra of the spectra cluster.

Note: Mesophyll and vascular bundle (cluster 4), epidermis (cluster 1, 2, 3).

Figure 5. Overlay of the second derivative of the average spectra from a cassava leaf (control) compared with CaSUT007 treated plants. (a) Epidermis and (b) Mesophyll.

Note: Average second derivative spectra after 13 points of smoothing and normalization with extended multiplicative signal correction over the spectral range of 1800–900 cm^–1.

Table 1. Band assignment of IR spectra presented in the plant tissues.

Frequency range (cm^−1)	Functional groups	References
1730	C=O ester from lignin and hemicellulose/Pectin	Kenneth and Lawrence (2005) and Dokken and Davis (2007) and McCann et al. (1992)
1650	Amide I band (C=O Stretch (80%), C–N stretch (10%), N–H bending (10%)	Yu (2004, 2005, 2008) and Pelton and McLean (2000), Nelson (1991) and Wetzel and Levine (2000)
1550	Amide II (N–H bend (60%), C–N stretch (40%)	Yu (2004, 2005) and Pelton and McLean (2000); Nelson (1991); Wetzel and LeVine (2000)
1510	C=C aromatic ring from lignin	Colthup et al. (1964), Campbell and Sederoff (1996) and Wetzel and LeVine (2000)
1450	Asymmetric CH2 and CH3 bending from lipids, protein, lignin	Kenneth and Lawrence (2005) and Colthup et al. (1964)
1370	Symmetric CH2 and CH3 bending from lipids, protein, lignin	Kenneth and Lawrence (2005) and Colthup et al. (1964)
1246	C–O Stretching from hemicelluloses and lignin	Dokken and Davis (2007), Faix (1992) and Kenneth and Lawrence (2005)
1200–1000	Mainly C–O–C of polysaccharides; very complex and depends uponcontributions from polysaccharides, cellulose, hemicellulose, and pectins	Kacuráková et al. (2000) and Cael et al. (1975)
1105	C–O–C glycosidic ether mainly hemicellulose	Dokken and Davis (2007) and Wang et al. (2012)
1155	C–C ring cellulose	Dokken and Davis (2007) and Wang et al. (2012)
1022,1047,1080	C–C, C–O Stretching from carbohydrates	Dokken and Davis (2007) and Wang et al. (2012)

Figure 6. PCA analysis of a cassava leaf. (a) Score and (b) loading plots from PCA analysis of epidermis control groups compared CaSUT007 treated plants. (c) Score and (d) loading plots from PCA analysis of mesophyll control groups compared CaSUT007 treated plants.

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