*Forests* **2020**, *11*, 1253 *Forests* **2020**, *11*, x FOR PEER REVIEW 8 of 13

**Figure 4.** Mean number (± SEM) of male adults/per trap/per week of *Thaumetopoea pityocampa* collected with pheromone-baited traps, and mean number (± SEM) of nests/tree recorded in plots where mating disruption (MD) was applied compared to control plots in 2016 (top) and 2017 (bottom). Bars with different letters are significantly different, according to the paired *t*-test (*p* < 0.05). **Figure 4.** Mean number (±SEM) of male adults/per trap/per week of *Thaumetopoea pityocampa* collected with pheromone-baited traps, and mean number (±SEM) of nests/tree recorded in plots where mating disruption (MD) was applied compared to control plots in 2016 (top) and 2017 (bottom). Bars with different letters are significantly different, according to the paired *t*-test (*p* < 0.05).

### **4. Discussion**

Although *T. pityocampa* is a key pest of pine trees in the Alps, there is little information about its phenology and seasonal occurrence in the north-western Italian Alps. In the Aosta Valley, PPM infestation has been consistently investigated in forest monitoring programmes from the 1960s onwards, but the defoliator moth has only been intensively studied since the 2000s. The Aosta Valley pine forests mainly consist of black pine and Scots pine; defoliation damage ranging from mild to very severe, due to PPM feeding activity, has been observed in about 50% of this area over the years (IR and LD). In 2016, the Department of Agriculture and Natural Resources, therefore, activated a plan to control PPM outbreaks.

Generally, mixed forests are less sensitive to pests and disease than pure stands. Although olfactory cues are commonly used to locate host trees, visual cues may affect the degree of infestation as well. Even in the case of PPM, mixed pine stands have been found to suffer less damage than pure pine stands [21], as non-host trees may act as physical barriers [22,23]. However, over the last century, several species of the genus *Pinus* have been widely used for afforestation [24], and this has clearly facilitated the spread of the PPM in the surveyed area. *P. nigra* is considered the primary native host of PPM, and the planting of this species has almost certainly promoted PPM population growth and outbreaks. Furthermore, PPM is reported to be sensitive to the severity and duration of the cold winter period, since it overwinters at the larval stage. The lethal temperature threshold is between approximately −10 ◦C and −16 ◦C [3], and the mean temperature increase over the last few decades has, thus, favored a higher survival rate from year to year, as already reported by Robinet [25]. In Aosta Valley, Mont Avic was used as a study site to monitor the expansion of PPM in the Italian Alps, where both an elevation gain (113.7 ± 23.0 m) and distance gain (232.7 ± 22.4 m) were observed in 2003 [26].

In our survey area, the network of permanent plots established in 2015 revealed that the PPM population progressively increased to reach its highest values in 2017, and then decreased rapidly afterward. The seasonal flight activity started at the beginning of June and continued until mid-September. The trapping of male adults in pheromone-baited traps peaked between the end of June and the beginning of July, with a similar trend in each year that was consistent with the literature [27,28]. Reliable population density data from monitoring and assessment surveys are required for effective suppression of PPM infestations. Although the absolute number of catches per trap may not be entirely representative of the population, being strongly influenced by sex ratio and trap frequency and position [24], monitoring traps may be useful to identify low- or high-density population areas where physical removal of the nests, insecticides, microbial treatments and/or mating disruption can be applied. Since several control methods are available, a practical evaluation of their effectiveness is essential for decision making in current PPM control protocols.

The Aosta Valley is a renowned tourist area, and in the past decades, there has been an increasing need for effective pest management programs, with traditional control methods (e.g., intensive pruning or chemical pesticides) strongly discouraged. The most common environmentally friendly control strategies (e.g., mechanical, biological) have now been adopted in the forestry environment.

Microbial insecticides offer a great deal of promise for pest management, being effective in controlling several defoliating forestry insect pests (e.g., spruce budworm, gypsy moth), without posing the serious environmental hazards associated with conventional pesticides [29,30]. Several microbial antagonists (e.g., bacteria, fungi, and viruses) have been reported for the control of PPM infestations [16], but the only large database based on scientific investigations concerns the use of *Bacillus*-based preparations. Applications of commercially available bioinsecticide products based on *Btk* have increased in the last few decades, targeting PPM [16,31,32], and other forest-defoliating lepidopterans [33–35]. At the surveyed sites, different control strategies were applied with the aim of containing and mitigating PPM outbreaks. *Btk* applications and mating disruption showed promising results in terms of effectiveness in controlling the infestations. Where *Btk* treatment was performed, the PPM mortality was very high. The average larval mortality was 90.47% in the treated plots *versus* 1.56% in the control plots. Similar values, with PPM larval mortality rates up to 90%, have previously been reported for *Btk* ground application in EU and Mediterranean countries [16,32]. In the literature, the susceptibility of the larvae to *Btk* was found to be directly proportional to the application dose, and some authors reported that a second *Btk* spray within 10–15 days of the first application might be useful [16]. In the current study, only one application was performed targeting L1–L2 larvae, using the dose of 32,000 BIU ha−<sup>1</sup> , and the average larval mortality rate proved very high at all the surveyed sites and in all years, making a second application unnecessary. Similar to *Btk* applications, the use of pheromones to disrupt the mating of PPM was effective at all the surveyed sites, suggesting that pheromone lures may effectively decrease male captures. The number of males in pheromone-baited traps was significantly lower in all MD plots (on average, 11.47 males/captured/week in all sites) in comparison with the control (on average, 119.16 at all sites) during 2016–2017. The number of nests per tree also confirmed the effectiveness of this technique, as also demonstrated by Trematerra [36]. The values recorded were significantly lower in all MD plots (on average, 0.7 nests/tree) compared to the control plots (on average, 1.87 nests/tree), highlighting the success of this technique. Although some constraints regarding the size of the plot have been reported for agricultural pests because of the risk of adult immigration [37], our investigations were performed in 1-ha plots, and the satisfactory results are consistent with those reported elsewhere [36].

PPM has a typical cyclical pattern, where intermittent outbreaks are interspersed with periods of lower population densities. In the literature, peak defoliations are reported to occur every 6–11 years on average [38–40], while in the Italian Alps, no regular cycles have been detected [41] (although the different statistical methods applied may have influenced the final output). A negative gradation phase characterized by a decline in the PPM population commonly occurs when there is a significant reduction in food availability. In the current study, the PPM population experienced a clear decline in 2019, but given that pine stands were not severely defoliated, it is more likely that this was due to the effectiveness of the set of control strategies applied rather than to the PPM's cyclical pattern. The pest is a strong public-health concern, due to the presence of urticating hairs that are present from the L3 larvae onwards, and which may affect humans, pets, and livestock in urban areas and forestry [13]. *Btk* applications and mating disruption have turned out to be the most appropriate control strategies for forest managers targeting the PPM. The adoption of such control strategies should be strongly encouraged by local authorities and administration policies, especially in areas frequented by people and in pine stands used for biomass production. Repeated annual applications of microbials and/or mating disruption could dramatically reduce population densities to levels below the economic injury level, with populations subsequently maintained at low levels with little management effort [42].

The role played by natural enemies is currently being investigated in Europe [40] and some Italian regions [42,43], and at least 18 parasitoid and 15 predator species have been recorded on the different stages of PPM. To date, the establishment of natural enemies in the Aosta Valley has not been studied, and little is known about their presence and potential impact along latitudinal or longitudinal gradients. Some questions remain to be addressed, and further studies should focus on the occurrence and adaptation of invertebrate natural enemies regarding PPM population dynamics. Since it is not completely clear how natural enemies respond to PPM outbreaks and what their role is in population regulation, specific research is needed to deepen our knowledge about the potential contribution and beneficial role of generalist parasitoids in controlling this defoliator moth in the surveyed area.

Further studies must also be performed regarding climatic conditions. Changes in diel activity and seasonal phenology patterns of both the PPM and oak processionary moth have been associated with global climate change [44], and moth emergence may be strongly affected by weather conditions, as already reported for the congeneric species *T. wilkinsoni* Tams [45]. All these further investigations, in addition to the data already recorded about seasonal occurrence and effectiveness of control strategies, will help provide a current global picture of PPM in the Aosta Valley, and allow for effective and sustainable management strategies to be adopted in the long term.

**Supplementary Materials:** The following are available online at http://www.mdpi.com/1999-4907/11/12/1253/s1, Table S1: List of the monitored municipalities in Aosta Valley in 2015–2019, Table S2: Total number of male adults of *Thaumetopoea pityocampa* monitored with pheromone-baited traps in Aosta Valley in 2015–2019.

**Author Contributions:** Conceptualization, C.F., I.R., L.D.; methodology, I.R., L.D.; formal analysis, C.F., V.S., C.P., I.R.; data Curation, C.F., V.S., I.R.; writing—Original draft preparation, C.F., V.S.; writing—review & editing, C.P., I.R., L.D., F.V.; project administration, L.D., F.V. All authors contributed to the writing of the manuscript and approved the final version. All authors have read and agreed to the published version of the manuscript.

**Funding:** This project was partially funded by the "MONGEFITOFOR" Project Interreg Cooperation Program Va ITA-CH 2014/2020.

**Acknowledgments:** The authors would like to thank all the forestry technicians of "Corpo Forestale della Valle d'Aosta" (CFVDA), and the phytosanitary service of the Aosta Valley for performing the field activity; Mario Negro, and Pierre-Yves Oddone for gathering data and assistance, and Raffaele Zanchini for the support in statistical analysis. The authors would like to thank the anonymous reviewers for their input in helping to improve the manuscript.

**Conflicts of Interest:** The authors declare no conflict of interest.

### **References**


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