1. Introduction
The pathogenic fungus,
Caliciopsis pinea Peck that causes Caliciopsis canker, has been found infecting native and exotic pines in Eastern North America and in Europe. In Eastern USA,
C. pinea has been reported on native
Pinus L
.:
P. strobus,
P. echinata Mill,
P. pungens Lamb,
P. rigida Mill,
P. virginiana Mill, and
P. taeda L. [
1]. In Europe,
C. pinea has been observed on exotic
Pinus:
P. echinata,
P. pungens,
P. rigida,
P. virginiana, and
P. radiata D. Don [
1]. Caliciopsis canker has also been described in native European
Pinus:
P. pinaster Aiton
, P. pinea L., and
P. halepensis Mill [
2]. Other species within the genus
Caliciopsis infect conifers in Western North America [
3]. Trees of all age classes are affected and saplings are sometimes killed. External symptoms include profuse pitching and cankers in the main stem and branches (
Figure 1a,b). Ascocarps are stalked, look like black spines 1–3 mm long and can be observed at the surface of roughed bark at the axils of branches (
Figure 1c) [
4].
Caliciopsis pinea has been referred to as a weak pathogen of
P. pinaster in France [
5]. The pathogen, however, can cause extensive damage to
P. strobus (eastern white pine) in Northeastern USA where saplings are killed and the wood quality of saw timber can be degraded (
Figure 1d) [
4,
6,
7,
8,
9].
Pinus strobus is economically and ecologically important in New England.
Pinus strobus comprises the greatest portion of the growing stock in Massachusetts (24%) followed by New Hampshire (21%) as well as 10% of Maine’s growing stock [
10].
Pinus strobus represents 60% and 30% of what sawmills produce in New Hampshire and Maine, respectively.
Pinus strobus also provides unique habitat for wildlife that other large conifer species cannot duplicate [
11]. The current
P. strobus resource, however, is aging as regenerating stands are rare [
10].
Reports of damage caused by
C. pinea were common in the 1930s, when initial studies on the pathogen’s epidemiology were conducted [
4,
12]. Since then, there has been very little progress towards understanding the epidemiology of this pathogen. Recently significant damage has been reported [
6,
7,
8]. Damage by
C. pinea has been associated with dense stands, suppressed trees, and poor sites. Very little quantitative data, however, exist about Caliciopsis canker distribution and site conditions associated with disease. The objective of this study, therefore, was to assess the extent and severity of Caliciopsis canker in the region of Eastern North America with the greatest concentration of
P. strobus and to explore environmental conditions associated with disease to identify areas at risk of damage. To meet our objective a survey was conducted in Maine, New Hampshire, and Massachusetts to determine presence and symptoms of
C. pinea in
P. strobus stands.
Figure 1.
(a) Caliciopsis canker in the main stem of Pinus strobus; (b) Profuse pitching resulting from Caliciopsis canker; (c) Ascocarps of Caliciopsis pinea, each 1–3 mm in length; (d) saw timber damaged by C. pinea.
Figure 1.
(a) Caliciopsis canker in the main stem of Pinus strobus; (b) Profuse pitching resulting from Caliciopsis canker; (c) Ascocarps of Caliciopsis pinea, each 1–3 mm in length; (d) saw timber damaged by C. pinea.
3. Results and Discussion
Caliciopsis canker symptoms were more widespread than expected (
Figure 2). Caliciopsis canker symptoms were observed in 42 (72%) of the 58 sites surveyed. Of these, 33 had sufficient pine and Caliciopsis canker symptoms to establish transects with 100 trees (>12.7 cm dbh) per site. Thirty five percent of the 3258 trees in transects had Caliciopsis canker symptoms (ranged from 3 to 87 affected trees per site). According to FIA data, the area forested by >75%
P. strobus basal area in Maine, Massachusetts, and New Hampshire, is 145,232 hectares with a sampling error of 12.4% (Randall Morin, U.S. Forest Service Northern Research Station’s Forest Inventory and Analysis, personal communication). Of the 18 FIA plots surveyed, 14 (78%) had Caliciopsis canker symptoms, suggesting that 112,958 hectares in these three states could potentially be affected by Caliciopsis canker.
Caliciopsis pinea was found causing cankers and fruiting on
P. strobus regeneration in 32 (55%) sites. The fungus fruits readily in the branch axils of
P. strobus saplings.
Caliciopsis pinea fruiting bodies on infected woody tissues were collected from 27 sites with symptomatic pines or seedlings and used for morphological and molecular identification.
Figure 2.
Location of 58 sites sampled in Maine, Massachusetts, and New Hampshire during 2014 and the presence of Caliciopsis canker symptoms at those locations.
Figure 2.
Location of 58 sites sampled in Maine, Massachusetts, and New Hampshire during 2014 and the presence of Caliciopsis canker symptoms at those locations.
Isolates of
C. pinea were successfully cultured from ascospores ejected onto inverted PDA, as well as from infected wood tissue directly plated on PDA after surface sterilization. The six sequences deposited in GenBank represent the first ITS sequences of
C. pinea, and as these isolates were recovered from
P. strobus in the northeast, just as the type culture was, they should be considered the best available proxy of the type culture ITS sequence. This is also of importance for future phylogenetic studies of all
Caliciopsis species as
C. pinea is the type specimen for the genus
Caliciospsis. In our phylogenetic analysis, all isolates of
C. pinea from North America cluster together into a single group, and the two isolates from Italy cluster together in a second distinct group with strong bootstrap support (94%) for this division (
Figure 3). Preliminary analysis would indicate isolates from North America and Italy represent distinct species, but further analysis of other genes and additional isolates from Italy and Europe are needed to draw any definitive conclusions. The other species of
Caliciopsis clustered together in a second group, and were more closely related to each other than to
C. pinea (
Figure 3). However, there is strong bootstrap support (95%) for all the
Caliciopsis species representing a single lineage, distinct from species of the closest known genus
Corynelia.
Figure 3.
Neighbor-joining phylogenetic tree based on the ITS sequence of fungi isolated from Caliciopsis pinea fruiting bodies or infected plant tissue, as well as closely related species imported from Genbank or provided by collaborators. C. pinea isolates are followed by host species and location the isolate was recovered from. Remaining isolates are followed by their respective Genbank accession numbers. Bootstrap values >70% were included at the nodes.
Figure 3.
Neighbor-joining phylogenetic tree based on the ITS sequence of fungi isolated from Caliciopsis pinea fruiting bodies or infected plant tissue, as well as closely related species imported from Genbank or provided by collaborators. C. pinea isolates are followed by host species and location the isolate was recovered from. Remaining isolates are followed by their respective Genbank accession numbers. Bootstrap values >70% were included at the nodes.
Using the NIDRM
P. strobus host layer was an effective method to locate and randomly select
P. strobus stands to survey. For example, 20 out of 29 sites visited in Maine were indeed
P. strobus stands as predicted by NIDRM. Most sites surveyed were located in the northeastern coastal zone (32 sites), in the plant hardiness zone 5b with mean annual extreme minimum temperature ranging from −23° to −26 °C (30 sites) [
21], and on flat slopes (mean slope = 5%, range = 0%–20%). The majority of these nominal variables did not have a significant effect on the proportion of sites with Caliciopsis canker across their categories. For example, for the three EPA ecoregions: northeastern highlands, northeastern coastal zone, and Acadian plains and hills,
p = 0.46 indicating that the proportion of sites with Caliciopsis canker symptoms was independent of ecoregion. Similarly, the proportion of sites with Caliciopsis canker was independent of plant hardiness zone three categories: 4b–5a, 5b, and 6a–6b (
p = 0.28) and for the three slope categories: 0, 1%–9%, and 10%–20% (
p = 0.9).
Important forest soil groups, however, were a more promising predictor of the probability of a site having Caliciopsis canker symptoms compared to EPA ecoregions, plant hardiness zones, and slope. The proportion of sites with Caliciopsis canker symptoms was greater for excessively drained or poorly drained soils (soil groups IC, IIB, and NC) compared to well-drained more fertile soils (IA, IB, and IIA) (
Table 1) [
24]. The proportion of sites with Caliciopsis canker symptoms was independent of forest soil group at
p = 0.05, but significant at
p = 0.1. The proportion of trees with Caliciopsis canker symptoms was constant (34%–37%) across New Hampshire important forest soil groups (
Figure 4a). The severity of symptoms, however, varied among important forest soil groups (
p = 0.027) and was greater for the IC with excessively drained, nutrient poor soils than for the IA-IB-IIA groups with well-drained, more nutrient rich soils (
Figure 4b). Conceivably pines growing in better soils have more resources to grow and might be less susceptible to disease. If forest soils were indicative of Caliciopsis canker damage, it would be informative to foresters who are likely to use forest soils information to make management decisions [
22].
Table 1.
Proportion of sites with Caliciopsis canker symptoms across important forest soil groups.
Table 1.
Proportion of sites with Caliciopsis canker symptoms across important forest soil groups.
Soil Group | Description | Percentage Sites with Caliciopsis Canker Symptoms | No. Sites |
---|
IA-IB-IIA | well drained, loamy, favourable moisture conditions, fertility suitable for hardwoods | 59 | 27 |
IC | excessively drained, very dry, coarse textured soils derived from glacial outwash, fertility adequate for softwoods but not hardwoods | 86 | 21 |
IIB-NC | sites with poorly drained soils and low fertility | 78 | 9 |
Figure 4.
(a) Incidence; (b) and severity of Caliciopsis canker symptoms in relation to important forest soil groups. Bars represent means and standard errors. Values with the same letter are not statistically different (α = 0.05).
Figure 4.
(a) Incidence; (b) and severity of Caliciopsis canker symptoms in relation to important forest soil groups. Bars represent means and standard errors. Values with the same letter are not statistically different (α = 0.05).
Pinus strobus density was another promising predictive factor of the probability of a site having Caliciopsis canker symptoms.
Pinus strobus density for sites with Caliciopsis canker was 311 trees/ha compared to 220 trees/ha for sites without Caliciopsis canker (
p = 0.1). High stand density and shallow soils were associated with white pine decline in southern Maine where trees growing under these conditions were predisposed to drought as restricted rooting depth and competition from other trees resulted in less available water [
26]. Other stress factors including heavy loss of needles from any of several recently-occurring needle diseases (
Lophophacidium dooksii,
Lecanosticta acicola, and
Bifusella linearis), in addition to overstocking and drought may also facilitate infection by
C. pinea. To further explore the relationship between Caliciopsis canker incidence, soil and
P. strobus density studies are underway with more systematic sampling within each soil group and intensive stand density measurements at each site.
Tree size and crown class were good predictors of a tree having Caliciopsis canker symptoms. Pole size, suppressed, and intermediate trees were more likely to have Caliciopsis canker symptoms and more severe symptoms than sawtimber, codominant trees (
Figure 5 and
Figure 6). Sawtimber sized trees were more frequent (2481) than poletimber sized trees (776). Similarly, trees with codominant crown positions (2516) were more frequent than suppressed and intermediate trees (589).
Figure 5.
(a) Incidence; (b) and severity of Caliciopsis canker symptoms in relation to size class. Bars represent means and standard errors. Values with the same letter are not statistically different (α = 0.05).
Figure 5.
(a) Incidence; (b) and severity of Caliciopsis canker symptoms in relation to size class. Bars represent means and standard errors. Values with the same letter are not statistically different (α = 0.05).
Figure 6.
(a) Incidence; (b) and severity of Caliciopsis canker symptoms in relation to crown class. Bars represent means and standard errors. Values with the same letter are not statistically different (α = 0.05).
Figure 6.
(a) Incidence; (b) and severity of Caliciopsis canker symptoms in relation to crown class. Bars represent means and standard errors. Values with the same letter are not statistically different (α = 0.05).
Caliciopsis canker symptoms become increasingly difficult to diagnose with increasing tree size. Profuse pitching is indicative of recent infections and is relatively common in pole size trees with smooth bark. As trees increase in size and bark roughens, profuse pitching is less frequently encountered. Older cankers tend to be surrounded by blackened, rough, and sunken bark and may still have visible pitching, but it becomes more difficult to see as the resin darkens with time. Symptoms in affected stands, therefore, are more difficult to assess as trees grow from pole to timber size where trees without resinosis can exhibit internal damage. It is possible that we might have underestimated the number of affected trees as profuse pitching is a symptom of recent infections and trees with older, internal damage can lack external pitching. Alternatively, we might have overestimated the number of trees with Caliciopsis canker by relying on pitching as the main symptom associated with the disease, although we did exclude pitching associated with insects, mechanical damage, internal decay, and white pine blister rust. Given the difficulty of assessing internal damage caused by C. pinea from the ground, studies that include tree harvesting are underway to better quantify damage to the wood product and the reliability of pitching as a survey tool for this disease. Cankers of symptomatic trees are being assessed for the presence of C. pinea and other fungi.
Caliciopsis canker has been reported more frequently in the last decade [
6,
7]. Most
P. strobus stands in New England are characterized by high stocking which is associated with
C. pinea damage [
27]. Alternatively, damage by might have been underreported in the past when management of
P. strobus was dominated by the threat of white pine blister rust caused by the exotic pathogen
Cronartium ribicola [
28]. Symptoms of both diseases, such as resinosis, are similar and might have been misidentified. Management practices that favor white pine blister rust control might have favored Caliciopsis canker development. For example, high stand density was encouraged for white pine blister rust management to promote early branch pruning. Environmental conditions that precede Caliciopsis canker development are being investigated.