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Article

Bark Inclusions in Canes of Southern Highbush Blueberry and Their Impact on Cane Union Strength and Association with Botryosphaeria Stem Blight

by
Renee M. Holland
1,2,†,
Chris J. Peterson
3,
Philip F. Harmon
4,
Phillip M. Brannen
1 and
Harald Scherm
1,*
1
Department of Plant Pathology, University of Georgia, Athens, GA 30602, USA
2
Cooperative Extension, University of Georgia, Bacon County, Alma, GA 31510, USA
3
Department of Plant Biology, University of Georgia, Athens, GA 30602, USA
4
Department of Plant Pathology, University of Florida, Gainesville, FL 32611, USA
*
Author to whom correspondence should be addressed.
†
Current address: Holland Consulting & Research, Alma, GA 31510, USA.
Horticulturae 2022, 8(8), 733; https://doi.org/10.3390/horticulturae8080733
Submission received: 21 June 2022 / Revised: 3 August 2022 / Accepted: 10 August 2022 / Published: 15 August 2022
(This article belongs to the Section Biotic and Abiotic Stress)
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Abstract

:
Bark inclusions are an understudied structural defect in trees and shrubs. They consist of areas of bark on adjacent parts of stems or scaffolds, typically on the inner faces of a narrow fork, which become overgrown and internalized to occupy part of the wood between the stems. Here, bark inclusions are described for the first time to occur in cane unions at the crown of southern highbush blueberry (Vaccinium corymbosum interspecific hybrids) cultivars ‘Farthing’ and ‘Meadowlark’, both of which are characterized by a narrow, vase-shaped architecture at the base of the plant, leading to crowding of the canes. When affected canes were dissected at their bases, bark inclusions were visible internally as a line of compressed bark within the wood of adjoining canes, or as bark invaginations and fissures across part of or the entire cross-section of the cane. Externally, blueberry crowns with included bark were characterized by either an inward ridgeline of bark between canes of similar diameters emerging from the crown at a narrow angle from each other, or by the presence of girdling roots. Bark inclusions were observed in plants of all ages, from the nursery to mature production fields. The internal length of the bark inclusion correlated strongly with the external length of the inward stem bark ridgeline symptom as measured by destructive sampling in the field (r = 0.916, p < 0.0001, n = 20). When plants with and without bark inclusions were subjected to a winch test in the field, the probability of breakage for canes without included bark was significantly lower (p < 0.0002) than for those with included bark, and at the maximum applied force of 972.4 N, 95.2% of the canes with bark inclusions failed (i.e., broke at the crown), compared with only 52.6% for canes without included bark. In a survey across three fields, the number of bark inclusions per plant was significantly associated with an index of cane crowding (r = 0.286. p = 0.0267, n = 60), suggesting that plants with tight, crowded bases had more bark inclusions. In addition, there was a significant association (p < 0.0001) between the presence or length of bark inclusions and the intensity of Botryosphaeria stem blight in these fields. This study showed that bark inclusions occur commonly in certain southern highbush blueberry cultivars in the production conditions of Georgia and Florida, with negative implications for cane integrity and plant health.

1. Introduction

The shift toward increased use of mechanical harvesting in blueberry has led to a change in cultivars used commercially. For example, growers of southern highbush blueberry (Vaccinium corymbosum interspecific hybrids) in Georgia and Florida are increasingly planting machine-harvestable cultivars such as ‘Farthing’, ‘Meadowlark’, and ‘Legacy’ [1,2,3] as opposed to more traditional cultivars that require hand-harvesting to ensure adequate fruit quality and minimize losses [4]. The former cultivars are characterized by a plant architecture more suitable for mechanical harvest, which typically includes a narrow, vase-shaped base to allow for better contact between the canes and the harvester fruit catcher pans resulting in reduced fruit ground losses [5]. In cultivated highbush blueberry, the base of the plant consists of multiple (up to ten or more) canes of different diameters that emerge from a common crown and from which the branches and shoots emerge to form the shrub. In recent years, we have observed structural defects at the crown of some of these cultivars where the canes emerge and a possible association with increased susceptibility to breakage and to infection with Botryosphaeria stem blight, in some cases requiring replant of one-third of a field or even an entire field. Dissection of these crowns revealed bark inclusions, i.e., areas of bark on adjacent parts of stems or scaffolds which became overgrown to occupy the wood of the internal joint. Such bark-included junctions are a common but understudied structural defect that occurs in many woody species [6] and can result in branch failure [7]. Based on preliminary observations in blueberry plantings in Georgia and Florida, we hypothesize that bark inclusions are more common in plants where canes are crowded; such situations are common in cultivars with narrow bases, especially where two canes of similar diameter emerge from the same location at the crown and compete for dominance at the base of the plant. As the blueberry plant ages and grows, the two canes continue to grow together, and the mechanical pressure created from the codominant canes can cause the formation of included bark at their union. It is well-known that infection by the fungal complex causing Botryosphaeria stem blight of blueberry is favored by wounding or mechanical injury [8,9,10,11,12]. Thus, tissue weakened by bark inclusions could serve as a previously overlooked infection court for stem blight fungi.
Bark inclusions and their impacts on the integrity of branch junctions have been studied mostly in amenity trees in landscapes and urban environments. For example, questionnaire responses from arborists and tree experts in the United Kingdom identified that normally formed branch unions rarely failed but bark-included junctions failed frequently [6]. In an experimental example, mechanical testing of branch unions of red maple trees (Acer rubrum) by pulling with a winch revealed that those with included bark were about 20% weaker than those that were normally formed [13].
The occurrence of bark inclusions at the crown of blueberry plants has not been described in detail previously, nor has the effect of included bark on cane union integrity or its association with Botryosphaeria stem blight been quantified. The objectives of this study were, therefore, to: (1) Provide a first description of bark inclusions at the crown of blueberry plants in relation to plant architectural properties; (2) elucidate the effect of these bark inclusions on the relative strength of affected blueberry cane unions; and (3) determine whether there is statistical evidence for the co-occurrence of bark inclusions and Botryosphaeria stem blight based on field survey data.

2. Materials and Methods

2.1. Field Sites

Observations and data on the occurrence and properties of bark inclusions were collected over a period of 4 years in five studies involving ten commercial blueberry plantings in southeastern Georgia and central Florida (Table 1). Apart from study 4, which involved potted plants in a blueberry nursery, all plantings were located on sandy or sandy-loam soils amended with pine bark and planted to southern highbush blueberry cultivars ‘Farthing’ or ‘Meadowlark’, both of which are characterized by having a narrow base architecture suitable for machine-harvesting [14]. Plants were 3 to 7 years old at the time of study with spacing typically 0.6 to 0.9 m within rows and 3.7 m between row centers.

2.2. Characterization of Bark Inclusions

At each site, 10 to 20 plants were examined for the presence of bark inclusions at their crowns using a combination of external (nondestructive) and internal (destructive) visual assessments. External evaluations consisted of a macroscopic examination of the anatomy and structure of the bark tissue on all canes and cane unions at the crown of the plant. In some cases, loose bark was scraped off with a knife to better view the anatomical characteristics of the cane union. Internal assessments were made by cutting the cane unions open with a pair of Corona Dual Cut carbon steel bypass loppers (Corona Tools, Corona, CA, USA) to initially separate the canes, and/or using a cordless 18-V DeWalt reciprocating saw (Stanley Black & Decker, Fort Mill, SC, USA) to cut through the crown of the plant at the location of the cane union. Images of external and internal symptoms were taken using an iPhone 10 with a 12-MP camera.
Blueberry cane unions with normally formed wood (Figure 1A–C) typically exhibit a raised callus between the canes that is visible externally upon removal of loose bark at the junction. No included bark was visible in the cross-section of such cane unions (Figure 1D). Externally, bark inclusions were tentatively identified by looking for two symptom types. One was characterized by an inward ridgeline of stem bark between canes of similar diameters emerging from the crown at a narrow angle to each other (Figure 1E,F). Internally, this ridgeline symptom was associated with a line of compressed bark within the wood of the adjoining canes (Figure 1G). The second external symptom type was characterized by the presence of girdling roots at the base of the plant (Figure 1H). The corresponding internal symptom consisted of bark invaginations and fissures across part of or the entire cross-section of the cane (Figure 1I). All studies described below were conducted on plants showing the more common ridgeline bark inclusion symptom.

2.3. Correlation between External and Internal Assessment of Bark Inclusions

This study was conducted to determine whether an external, nondestructive assessment can quantitatively predict the occurrence of bark inclusions as verified by internal assessment following destructive sampling (study 2 in Table 1). A total of 20 plants (7 years old at the time of the assessment) were examined externally and internally for bark inclusions. For the external measurement, the length of the ridgeline bark symptom as it could be observed on cane unions on the intact plant (with or without removal of loose bark) was measured with a woven tape measure to the nearest mm. Then, after the canes had been separated and cut open as described above, the internal total length of the bark inclusion was measured similarly. Linear regression analysis in SigmaPlot v.11.0 (Sigma-stat Software, Chicago, IL, USA) was applied to determine the strength and significance of the correlation between internal and external measurement of bark inclusion length. t-tests were applied to determine whether the intercept and slope of the relationship were significantly different from 0 and 1.0, respectively.

2.4. Effect of Bark Inclusions on Cane Strength

Two 3-year old blueberry plantings on sandy loam soil were utilized in this study (study 5 in Table 1). At each site, ten plants having bark inclusions at the cane union (identified by external assessment) were selected, along with ten control plants having normally formed wood at their unions. Care was taken to select plants and test canes for uniformity with cane diameters typically 2 to 3 cm at 25 cm above ground level. The length of the externally visible ridgeline symptom was measured nondestructively as described above and recorded.
A static winching test following Peterson and Claassen [15] was conducted to determine the pulling force at the time of cane union breakage, with adjustments for the smaller size of blueberry bushes compared with the tree species evaluated by these authors. The cane from each plant selected for winching was in a straight, upright orientation with no other canes interfering with the path of the winching test. A nylon strap was wrapped around the cane at the transition point between lignified and green tissue but below the region where the cane was excessively branching to produce branches and shoots. The strap was attached to a Mark-10 Series 3 M3-200 digital force gauge (Mark-10, Copiague, NY, USA) rated for 90.7 kg (200 lbs) and was attached to an electrical winch on an all-terrain utility vehicle parked in a row middle ~1.8 m from the test plant. The angle of the strap from the horizontal was typically between −3 and −4° when the strap was tightened. The winch was engaged gradually to increase the pulling force on the cane, and the peak force reading from the force gauge was recorded by the instrument at the time of cane union failure (breakage) or when the cane was bent fully to the ground in the case of canes that did not break. The angle of the cane from the vertical at the time of breakage was recorded by visual observation with a 30.5-cm high-visibility laser-etched aluminum rafter square (Empire Level Manufacturing, Mukwonago, WI, USA).
Following the winch tests, each test cane, regardless of whether it broke or not, was dissected at its base to confirm the presence of a bark inclusion based on internal assessment. Failure time analysis (survival analysis) was applied to determine whether the survival probability distributions differed between canes with and without bark inclusions in relation to the force applied. For each of n = 40 plants, the dataset for the analysis consisted of (1) two binary variables (0 or 1) for each plant indicating whether the cane had an internal bark inclusion and whether the cane broke during the winch test; and (2) measurements of the force and angle from the vertical at which the cane union broke. Cane unions that did not break at the maximum force encountered during the winch tests (972.4 N) were considered censored observations in the failure time analysis (n = 10). The analysis was conducted in PROC LIFETEST is SAS 9.4 (SAS Institute, Cary, NC, USA) following Scherm and Ojiambo [16].

2.5. Associations between Bark Inclusions and Botryosphaeria Stem Blight

This study included three fields selected because symptoms of both bark inclusions and Botryosphaeria stem blight were present based on initial visual observations (study 3 in Table 1). Diseased cane cross-sections can be distinguished by having a brown, necrotic appearance compared with the textured, compressed bark layer from the bark inclusion (Figure 1J,K).
In each field, 20 plants were selected systematically to represent a range of Botryosphaeria stem blight severity classes (asymptomatic, light symptoms, moderate symptoms, and severe symptoms) with five plants from each class being assessed (Supplementary Figure S1). Asymptomatic plants did not show visually apparent stem blight symptoms. Light symptoms were characterized by one or two blighted canes per plant or “flagging” with most of the canopy remaining green and asymptomatic. Moderate symptoms showed two or more blighted canes with one-third to one-half green canopy, whereas severely symptomatic plants displayed general plant decline with several blighted canes and most of the canopy necrotic with only <20% of green canopy remaining. Stem blight severity for each plant was quantified by measuring the lengths of all the canes per plant and then expressing the length of the symptomatic portions of the diseased canes as a percentage.
On the same 20 plants per field, bark inclusions were identified visually as described above and their lengths measured externally using a tape measure. In addition, an index of cane crowding (in units of cm−2) was calculated for each plant as the number of canes divided by the cross-sectional area of the shrub at 25 cm above the ground. The cross-sectional area was derived from two diameter measurements perpendicular to one another of the whole plant base at 25 cm height. Large values of the cane crowding index indicate that more canes occupy the same plant space.
Pearson’s correlation coefficients were used to measure the strength and significance of the associations among the four measured variables: number of bark inclusions/plant, length of bark inclusions/plant, cane crowding index, and stem blight severity (%).
To confirm infection of plants by members of the Botryosphaeriaceae, symptomatic stem sections were taken from each field site, samples were excised from the stems, immersed in a 5% household bleach solution (0.25% sodium hypochlorite) for 3 min, rinsed with sterile distilled water, dipped into 95% ethanol and flame-sterilized, and plated onto one-fifth strength potato dextrose agar amended with 0.01 mg rifampicin and 0.25 g anhydrous ampicillin. The resulting fungal growth was transferred to acidified potato dextrose agar and sent to the Plant Molecular Diagnostics Lab at the University of Georgia Tifton campus for genus-level identification using primers ITS1F and ITS4 to amplify the internal transcribed spacer (ITS) region [17]. Identifications were based on BLAST comparisons with nucleotide sequences available in NCBI GenBank using default parameters, selecting the short queries option, and setting word size to 28.

3. Results

3.1. Occurrence of Bark Inclusions

Bark inclusions were observed commonly across the five studies, with the incidence of affected plants ranging from approximately 15 to 50%. The inward ridgeline of stem bark between two canes of similar diameters was the symptom that occurred more commonly than the other external symptom of girdling roots at the base of the plant. However, both symptoms co-occurred in the same field. Symptoms were observed on plants of all ages, from young plants in the nursery (Figure 2) to fully mature plants in the field. Furthermore, bark inclusions could be observed any time of the year but were more easily visible in the fall and during the dormant season due to lower weed density, removal of root sprouts at the base of the plant, and more open plant canopies due to leaf drop.

3.2. Correlation between External and Internal Assessment of Bark Inclusions

Linear regression analysis documented a strong correlation (r = 0.916, p < 0.0001, n = 20) between bark inclusion lengths as measured externally versus internally on Farthing southern highbush blueberry in the field (Figure 3). This validated the use of the external, nondestructive assessment for use in the remaining field studies in this paper. The intercept of the regression line was not significantly different from 0 (p = 0.788), whereas its slope (1.281 ± 0.077 [SE]) was significantly greater than 1.0, indicating that internal bark inclusions were slightly longer than suggested by the external measurement.

3.3. Effect of Bark Inclusions on Cane Strength

Blueberry canes with included bark broke at lower tension forces than those without included bark (Figure 4). Failure time analysis of the combined data set (two sites) showed that survival probabilities for canes without included bark were significantly higher (p < 0.0002 based on both log-rank and Wilcoxon test statistics) than those with included bark. At the maximum applied force of 972.4 N, 95.2% of the canes with bark inclusions failed (i.e., broke at the crown), compared with only 52.6% for canes without included bark (Figure 4).

3.4. Association between Bark Inclusions and Botryosphaeria Stem Blight

Across the three fields surveyed as part of study 3 (n = 60 plants), significant associations among bark inclusion, stem blight, and plant architecture variables were revealed (Table 2). Not surprisingly, the two bark inclusion variables (number and external length of bark inclusions) were strongly correlated with each other (r = 0.886, p < 0.0001). In addition, the number of bark inclusions per plant was also significantly associated with the cane crowding index (r = 0.286. p = 0.0267), suggesting that plants with crowded bases had more bark inclusions. However, stem blight severity was not significantly associated with the cane crowding index (p = 0.433). Both bark inclusion variables correlated significantly (p < 0.0001) with stem blight severity, with correlation coefficients of 0.488 and 0.515 for the two combinations (Table 2). The results thus document an association between the presence or intensity of bark inclusions and the severity of stem blight.
Isolations from the margins of stem blight symptomatic tissues yielded primarily fungi within the Botryosphaeriaceae (Lasiodiplodia and Neofusicoccum spp.) (Table 3), which are known as common causes of stem blight of blueberry in the southeastern United States [18]. In addition, members of the Pestalotiopsidaceae (Neopestalotiopsis and Pestalotiopsis spp.) and Trichosphaeriaceae (Nigrospora sp.) were also found. Neopestalotiopsis and Pestalotiopsis species are stem blight and dieback pathogens on blueberry in Peru and on other woody hosts, such as macadamia nut [19,20]. Nigrospora is typically a leaf spot pathogen but has also been associated with woody hosts, such as bamboo and Chinese rose [21]. Some isolations also yielded Trichoderma and Fusarium species which were considered secondary in the context of this study.
Although the species names of GenBank matches are shown in Table 3 for completeness sake, we acknowledge that the single-locus ITS analysis applied here only allows us to draw conclusions about the family or genus level of the isolated organisms. This is consistent with our overall goal of confirming infection of symptomatic canes with known stem blight fungal groups.

4. Discussion

This study showed that bark inclusions occur on certain southern highbush blueberry cultivars in the production and crop management conditions of southern Georgia and central Florida. Symptoms were readily observed on cultivars ‘Farthing’ and, to a lesser degree, on ‘Meadowlark’, both of which are characterized by a narrow base of the plant and/or an upright plant growth architecture to facilitate mechanical harvesting. Other cultivars were rarely affected, suggesting a genetic component to the propensity of forming bark inclusions. Our study revealed two external symptom types corresponding to bark inclusions, one characterized by an inward ridgeline of stem bark layers formed at a narrow fork on two adjoining canes, and the other by the presence of girdling roots at the base of the plant. Bark inclusions were found in plants of all ages, in some cases as early as in 1-year-old potted plants in the nursery. Potential causes leading to the two types of bark inclusions in blueberry are discussed below.
Bark inclusions of the inward ridgeline type have been described in the arboricultural literature [7,22]. They are caused by a plant response when two stems, canes, or branches emerge at a narrow fork, have diameters of similar sizes, and compete for dominance. The two canes press against each other, which causes the bark at the cane union to become wedged between the canes, resulting in enclosed bark. This weakens the cane union and predisposes it to splitting [23], reducing mechanical strength and forming a potential entry point for pathogens. The dependence of this type of bark inclusion on the presence of narrow, V-shaped forks is consistent with its occurrence in blueberry cultivars bred for a narrow base and upright plant architecture, such as ‘Farthing’ and ‘Meadowlark’ examined here.
The second type of bark inclusion observed in the present study, characterized externally by the presence of girdling roots at the base of the plant, is likely caused by poor plant management. When excessive nitrogen is used, the roots and crown of the blueberry plant can become twisted or constricted. This is of particular concern in nurseries, where root growth may be constrained by the container size, whether it be square trays, narrow plugs, or trade-gallon pots. This is of particular concern when planting schedules are delayed but plants continue to be fertilized and watered at high levels while still in their original containers. The resulting bound roots may not loosen after planting, and girdling or deformed roots may not recover after planting [24,25], especially when care is not taken to loosen roots and soil and/or plants are placed in compacted soil [26,27]. Furthermore, the advent of fertigation has increased the temptation to overfertilize young plants to enable rapid growth and early fruit production, but in such cases, the soil around the crown and roots may become so saturated that roots are unable to grow and become established [28,29]. These cultural malpractices applied to young plants in the nursery or during field establishment may result in the girdling roots symptom and associated internal bark inclusions observed here.
Further investigation of bark inclusions of the ridgeline type showed a close correlation between the bark inclusion length measured externally versus internally, allowing the external assessment to be used to assess the plants nondestructively. This is the first study to establish a method for assessing bark inclusions in blueberry and could be a useful tool for cultivar evaluation in blueberry breeding programs. Bark inclusions have been studied more extensively in shade trees and urban forestry, and similar techniques were employed to measure bark inclusions from the apex of a junction down to the base of the included bark [7]. Because of the convoluted pattern of bark inclusions in plants with girdling roots, typically involving multiple internal lobes and invaginations, no attempt was made to establish a relationship with internal bark inclusion length for this case.
Our winching study clearly showed that bark inclusions increase the risk of cane failure in blueberry, whereby cane unions with bark inclusions required significantly less force to break compared with those without this defect. Thus, the presence of included bark may predispose plants to cane breakage in response to mechanical forces encountered in the field, such as during machine-harvesting or in response to high winds associated with severe weather, tropical storms, or hurricanes, which can occur in the southeastern United States [10,13,30,31,32]. Winter injury may be another predisposing factor to cane union failure in the presence of bark inclusions. In rhododendron shrubs, winter injury can occur by water getting trapped in bark inclusions in narrow crotch angles [33]. The water freezes and expands, which in turn forces the canes apart at the union creating a wound.
Most of our studies were conducted on plants showing no symptoms of disease. However, it is well-known that plant stress and injuries at the crown and on lower canes can predispose blueberry plants to stem blight, caused primarily by fungal members of the Botryosphaeriaceae [10,34]. Stem blight occurs commonly in the southeastern United States, and in Georgia and Florida is caused predominantly by the species Lasiodiplodia theobromae and Neofusicoccum ribis with occasional isolates of Botryosphaeria dothidea and Diplodia seriata [18]. Based on stem blight field surveys in three fields, we document here for the first time an association between stem blight intensity and the presence of bark inclusions. Common stem blight fungal groups known to be pathogenic on blueberry were readily isolated from the diseased canes in these surveys, although the limited resolution of the ITS analyses applied here did not allow us to draw conclusions about the exact species present. Although further research is needed to establish a cause-effect relationship between bark inclusions and increased susceptibility to stem blight, e.g., by artificial inoculation of plants with and without bark inclusions, this survey provides the first empirical evidence for the common occurrence of bark inclusions in certain blueberry cultivars and their detrimental effect on cane integrity.
There are several potential means to mitigate bark inclusions in blueberry. For inclusions with the bark ridgeline symptom, cane pruning may be applied to reduce cane crowding but also to allow for canes of different diameters to lessen the issue of codominance and eliminate tight, narrow junctions. This would allow for normal wood formation at cane junctions. For the girdling roots symptom, proper horticultural practices in the nursery and upon plant establishment could help alleviate this symptom. Nurserymen and growers alike should be aware that allowing plants to become pot-bound, either in the nursery, after plant order receipt, or in response to delays in planting at the field site may predispose plants to the aforementioned girdling roots symptom. This symptom may also be exacerbated by not properly loosening roots at planting, planting in compacted soil, or overly aggressive fertility programs.

5. Conclusions

Bark inclusions, a structural defect at the crown of the plant, are described for the first time in southern highbush blueberry, and a correlation between external and internal assessments of bark inclusions is presented. Canes with included bark broke at lower tension forces than those without, documenting a significant negative impact on cane strength. The number of bark inclusions per plant was associated with cane crowding, and bark inclusion intensity correlated with Botryosphaeria stem blight severity in field surveys. Horticultural practices to mitigate the incidence and impact of bark inclusions are discussed.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/horticulturae8080733/s1, Figure S1: Botryosphaeria stem blight severity classes used to pre-select plants in the stem blight survey in relation to bark inclusions: asymptomatic (A), light (B), moderate (C), and severe (D).

Author Contributions

R.M.H.—conceptualization, methodology, formal analysis, investigation, data curation, writing (original draft); C.J.P.—methodology, writing (review and editing); P.F.H.—conceptualization, investigation, writing (review and editing); P.M.B.—resources, writing (review and editing), supervision; H.S.—conceptualization, methodology, investigation, resources, writing (review and editing), supervision, project administration, funding acquisition. All authors have read and agreed to the published version of the manuscript.

Funding

Supported by Hatch Act and State of Georgia funds.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

The raw data underlying these analyses are available from the corresponding author upon reasonable request.

Acknowledgments

We thank the blueberry growers who allowed us to conduct the surveys and experiments reported here in their fields. Assistance by grounds foreman Shawn Thompson and plant diagnostician Ansuya Jogi is gratefully acknowledged.

Conflicts of Interest

The authors declare no conflict of interest.

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Figure 1. Anatomy of normally formed southern highbush cane unions (A–D) compared with those having included bark (E–K). External view of a normally formed cane union prior to (A) and after removal of loose bark (B,C). Note raised line of callus between the two canes in (C), with no bark inclusion visible internally in the cross-section of the cane union (D). This is compared with a narrow, V-shaped cane union with included bark that can be viewed externally by an inward ridgeline bark layer (E,F) and internally as compressed bark in the cane union cross-section (G). Another symptom of bark inclusions can be identified externally as girdling roots at the basal crown (H) and internally as bark fissures across the cane cross-section or as a partial invagination on one side of the cane (I). Diseased, necrotic cane tissue due to stem blight (J) can be differentiated from dark, compressed bark inclusion line (K). Blue brackets or lines indicate length of included bark or bark fissure.
Figure 1. Anatomy of normally formed southern highbush cane unions (A–D) compared with those having included bark (E–K). External view of a normally formed cane union prior to (A) and after removal of loose bark (B,C). Note raised line of callus between the two canes in (C), with no bark inclusion visible internally in the cross-section of the cane union (D). This is compared with a narrow, V-shaped cane union with included bark that can be viewed externally by an inward ridgeline bark layer (E,F) and internally as compressed bark in the cane union cross-section (G). Another symptom of bark inclusions can be identified externally as girdling roots at the basal crown (H) and internally as bark fissures across the cane cross-section or as a partial invagination on one side of the cane (I). Diseased, necrotic cane tissue due to stem blight (J) can be differentiated from dark, compressed bark inclusion line (K). Blue brackets or lines indicate length of included bark or bark fissure.
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Figure 2. Bark inclusions in trade-gallon potted nursery plants, external (A), internal partial (B), internal total (C). Blue line shows length of external bark inclusion (ridgeline type).
Figure 2. Bark inclusions in trade-gallon potted nursery plants, external (A), internal partial (B), internal total (C). Blue line shows length of external bark inclusion (ridgeline type).
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Figure 3. Linear regression analysis of the internal assessment of bark inclusion length vs. the external (nondestructive) assessment of bark inclusion length in canes of Farthing southern highbush blueberry (n = 20, study 2). The two measures were strongly correlated (r = 0.916, p < 0.0001), and the slope of the regression line was 1.281 ± 0.077 (SE). The dashed line corresponds to a 1:1 relationship.
Figure 3. Linear regression analysis of the internal assessment of bark inclusion length vs. the external (nondestructive) assessment of bark inclusion length in canes of Farthing southern highbush blueberry (n = 20, study 2). The two measures were strongly correlated (r = 0.916, p < 0.0001), and the slope of the regression line was 1.281 ± 0.077 (SE). The dashed line corresponds to a 1:1 relationship.
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Figure 4. Product limit survival estimates of Farthing southern highbush blueberry cane unions with or without included bark in relation to tension applied to the cane with a winch (n = 40, study 5). Survival probability in this context refers to the proportion of canes that did not break at a given applied force. Censored data refer to canes that did not break at the maximum applied force of 972.4 N.
Figure 4. Product limit survival estimates of Farthing southern highbush blueberry cane unions with or without included bark in relation to tension applied to the cane with a winch (n = 40, study 5). Survival probability in this context refers to the proportion of canes that did not break at a given applied force. Censored data refer to canes that did not break at the maximum applied force of 972.4 N.
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Table
Table 1. Field studies in which bark inclusion at cane junctions of southern highbush blueberry plants were investigated.
Table 1. Field studies in which bark inclusion at cane junctions of southern highbush blueberry plants were investigated.
Study No.ActivityLocation and YearNumber of FieldsCultivar
1Initial characterization of bark inclusionsGainesville, FL, USA (2017)3‘Farthing’, ‘Meadowlark’
2External versus internal assessment of bark inclusionsAlma, GA, USA
(2017)		1‘Farthing’
3Stem blight survey in relation to bark inclusionsHomerville, GA, USA
(2018)		3‘Farthing’
4Examination of bark inclusions in a blueberry nurseryAlma, GA, USA
(2019)		1‘Farthing’
5Effect of bark inclusion on cane strengthAlma, GA, USA
(2020)		2‘Farthing’
Table
Table 2. Pearson’s correlation coefficients and associated p-values (in parentheses) showing the associations among bark inclusion, Botryosphaeria stem blight, and plant architecture variables in Farthing southern highbush blueberry (n = 60, study 3).
Table 2. Pearson’s correlation coefficients and associated p-values (in parentheses) showing the associations among bark inclusion, Botryosphaeria stem blight, and plant architecture variables in Farthing southern highbush blueberry (n = 60, study 3).
Number of Bark Inclusions/PlantLength of Bark Inclusions/Plant (cm)Stem Blight Severity (%)Cane Crowding Index (cm−2) a
Number of bark inclusions/plant1.0000.886
(<0.0001)		0.488
(<0.0001)		0.286
(0.026)
Length of bark inclusions/plant (cm) 1.0000.515
(<0.0001)		0.209
(0.107)
Stem blight severity (%) 1.0000.103
(0.433)
Cane crowding index (cm−2) a 1.000
a Number of canes divided by cross-sectional area of plant at 25 cm height.
Table
Table 3. BLAST results from ITS sequence comparisons showing example accessions that were highly similar to our isolate sequences obtained from southern highbush blueberry canes with bark inclusions and stem blight symptoms in study 3.
Table 3. BLAST results from ITS sequence comparisons showing example accessions that were highly similar to our isolate sequences obtained from southern highbush blueberry canes with bark inclusions and stem blight symptoms in study 3.
SiteIsolateBest Matches (Percent Identity) aQuery Coverage (%) a
11Neopestalotiopsis clavispora KR052094.1 (100), N. rosae (99.8), Pestalotiopsis microspora (99.8)100
12Lasiodiplodia mediterranea KU578251.1 (100), L. viticola (100), L. theobromae (99.7)100
14Neofusicoccum parvum KU997486.1 (100), N. kwambonambiense (99.8)96
18Neopestalotiopsis clavispora MT240541.1 (100), Pestalotiopsis clavispora (100), N. rosae (99.8)100
116Neofusicoccum parvum MW532988.1 (100), N. kwambonambiense (100), N. umdonicola (99.8)99
118Neofusicoccum parvum MH779823.1 (100), N. kwambonambiense (99.8), N. umdonicola (99.8)100
23Neopestalotiopsis clavispora MH429984.1 (100), Pestalotiopsis microspora (100), P. clavispora (100)100
26Pestalotiopsis sp. KM520039.1 (100), P. microspora (99.8), Neopestalotiopsis clavispora (99.8)100
211Pestalotiopsis microspora KM438014.1 (100), P. clavispora (100), Neopestalotiopsis saprophytica (99.8), N. clavispora (99.8)100
217Pestalotiopsis sp. JN418796.1 (100), P. microspora (99.8), Neopestalotiopsis clavispora (99.6)100
220Lasiodiplodia theobromae MT302844.1 (100), L. citricola (100)100
222Nigrospora sphaerica KJ767121.1 (100), N. oryzae (99.4)99
35Neofusicoccum parvum MF449509.1 (100), N. kwambonambiense (100), N. umdonicola (100), N. ribis (100)100
313Neofusicoccum parvum KX648507.1 (100), Lasiodiplodia theobromae (99.6)100
314Neofusicoccum parvum JQ647908.1 (100), N. kwambonambiense (100), N. umdonicola (99.8)99
315Pestalotiopsis microspora MT071251.1 (100), Neopestalotiopsis clavispora (100)100
a GenBank number and percent query coverage provided for top match. Although ITS analysis alone is insufficient to identify most of these organisms to species level, the analysis confirmed the presence of known stem blight fungal groups (Botryosphaeriaceae, Pestalotiopsidaceae, and Trichosphaeriaceae).
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Holland, R.M.; Peterson, C.J.; Harmon, P.F.; Brannen, P.M.; Scherm, H. Bark Inclusions in Canes of Southern Highbush Blueberry and Their Impact on Cane Union Strength and Association with Botryosphaeria Stem Blight. Horticulturae 2022, 8, 733. https://doi.org/10.3390/horticulturae8080733

AMA Style

Holland RM, Peterson CJ, Harmon PF, Brannen PM, Scherm H. Bark Inclusions in Canes of Southern Highbush Blueberry and Their Impact on Cane Union Strength and Association with Botryosphaeria Stem Blight. Horticulturae. 2022; 8(8):733. https://doi.org/10.3390/horticulturae8080733

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Holland, Renee M., Chris J. Peterson, Philip F. Harmon, Phillip M. Brannen, and Harald Scherm. 2022. "Bark Inclusions in Canes of Southern Highbush Blueberry and Their Impact on Cane Union Strength and Association with Botryosphaeria Stem Blight" Horticulturae 8, no. 8: 733. https://doi.org/10.3390/horticulturae8080733

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