Introduction

The marsh fritillary Euphydryas aurinia has received much attention, especially in the UK and Europe, owing to its protected status in many countries, its inclusion on the EU Habitats Directive, and because it is declining rapidly. Most ecological research has focused on its conservation – for example its status (Warren 1994; Lewis and Hurford 1997; Hula et al. 2004; Fowles and Smith 2006), its habitat and how best to manage it (Anthes et al. 2003; Konvicka et al. 2003; Betzholtz et al. 2007; Smee et al. 2011; Botham et al. 2011), and how much and what to conserve (Wahlberg et al. 2002; Schtickzelle et al. 2005; Bulman et al. 2007). The densities of young caterpillars in the autumn, when they live communally in aggregations and construct conspicuous silk structures or ‘webs’ in which they feed and moult, is the unit by which status is measured. This is before caterpillars have negotiated winter hibernation, when mortality is highest (Liu et al. 2006), and undergone the majority of development in the following spring.

Invariably, an elevated density of autumn webs is correlated with higher cover of the foodplant devil’s bit scabious Succisa pratensis (Konvicka et al. 2003; Betzholtz et al. 2007; Smee et al. 2011; Botham et al. 2011; Pschera and Warren 2018) and foodplant cover is regarded as paramount (e.g. Fowles and Smith 2006; Porter and Ellis 2011). The biology of the caterpillars themselves has been less studied, especially post-hibernation ecology and development to pupation in the spring, aside from their relations with host-specific parasitoid wasps (Porter 1983; Klapwijk et al. 2010).

Populations of E. aurinia are volatile throughout the UK and Europe, crashing in some years before appearing again, often suddenly and in substantial numbers (Asher et al. 2001; Zimmerman et al. 2011). A survey of the status of E. aurinia on the Isle of Islay in Scotland was undertaken in spring 2012 during the early growth of post-hibernation caterpillars and during a phase of pronounced population contraction (Ravenscroft 2013). The species was extremely scarce and only one or two webs of caterpillars occurred at a few sites. After the flight period of 2012, caterpillars were absent in the autumn at some locations occupied in the spring, but they were recorded again at these locations in spring 2013. These caterpillars appeared unusually advanced, being larger and a later instar than caterpillars elsewhere.

Further study from 2013–2017 considered the implication of these ‘advanced’ caterpillars – that E. aurinia may overwinter more than once before pupating. Prolonged life-cycles occur in many insects in response to challenging environments that are temporary, dry, unstable or cold and are usually facilitated by suspension of one stage of growth, especially pupation (Powell 2001; Benyamini 2008; Saulich 2010). Many larvae of Lepidoptera in alpine or arctic regions or at the edges of their ranges have extended life-cycles (Wipking 1998; Fischer and Fiedler 2001; Pratt and Emmel 2010) as does E. aurinia in Sweden (Eliasson and Shaw 2003). Sister species from northern latitudes or at high altitude may also take two years to develop from egg to adult e.g. E. intermedia in the Alps (Luckens 1985), E. gillettii in the Rockies of North America (Williams et al. 1984) and E. maturna in Finland (Wahlberg 1998). The arctic E. iduna is believed to take several years to develop (Ehrlich and Hanski 2004). But there is no indication that E. aurinia has a longer life-cycle at low altitude in a relatively mild temperate zone such as the UK.

Studies on Islay concentrated initially on establishing the normal development of larvae in the field, from the early instars in autumn soon after hatching through to pupation in the following spring. Then they considered whether unusually advanced caterpillars in the early spring could be the product of an annual life cycle by investigating their occurrence and comparative biometrics, and they also investigated whether parasitism might be responsible for accelerated growth. Field experiments then manipulated the distribution of new generations of caterpillars, rendering areas free of these in spring in which the occurrence of caterpillars of advanced development would confirm an extended life-cycle for E. aurinia.

The life-cycle of E. aurinia

Adult E. aurinia fly primarily in June in Scotland and their eggs, laid in batches on the larval foodplant devil’s-bit scabious Succisa pratensis, hatch in July (Fig. 1). Caterpillars live and feed communally, hidden initially under the leaves of the foodplant until mid-August, but thereafter more visibly in characteristic tents of silk, or ‘webs’, that are spun over the foodplant. They undergo three moults together in the autumn before hibernating soon after reaching the 4th instar in September or October (occasionally late August). This instar spins balls of silk deep within the vegetation near ground level in which the entire web of caterpillars hibernates.

Fig. 1
figure 1

The annual life-cycle of E. aurinia in Scotland, showing the spans of larval instars. After hatching, early instars occur in the autumn, hibernate in the 4th instar, and emerge and complete development the following spring. The period of hibernation is shown in black

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Caterpillars emerge from hibernation in late February and March, and after spinning an initial web on which they bask together for a period, continue feeding before moulting to the 5th instar, usually in early-mid April. These remain social in small groups before the final moult to the 6th instar that is solitary and completes development to pupation in late April or May.

Throughout this paper, hibernation refers to the winter period of dormancy undertaken by 4th instar caterpillars between autumn and spring, and diapause refers to longer periods of dormancy that may be undertaken by caterpillars.

Methods

Study sites and status of E. aurinia

Islay is a large island, about 620 square kilometres in area and 20 miles or so from the west coast of Scotland. Transects were used to assess the presence and distribution of larval webs of E. aurinia at known locations for the species across the island in each autumn 2012 to 2016 (Fig. 2). Webs were recorded 1 m either side of measured walkabouts, or straightforward searches of smaller sites (ca < 0.5 ha), and their locations were recorded with a GPS and marked with white plastic posts. Transects were supplemented by searches if no webs were recorded.

Fig. 2
figure 2

The distribution of 1 km squares on Islay containing sites studied in autumn and spring 2012–2016

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Study sites were units of contiguous habitat that were easy to identify by features of the landscape, thus facilitating repeatability during surveys. They were generally less than 2 ha in extent and were usually part of extensive landscapes of patchy habitat. Most were in separate 1 km squares of the Ordnance Survey UK grid (Fig. 2) but those in the same square were separated from other sites by at least 100 m. There is so much habitat for E. aurinia on Islay, and few barriers to movement, that it is not practical to define boundaries or areas that might be considered isolated from others.

Identification of instars

Instars of E. aurinia were separated by overall appearance and size, but primarily on the basis of the head capsule that is very distinctive: this is about 0.75 mm wide in 3rd instars; 1 mm in 4th instars; 1.5 mm in 5th instars; and 2 mm in 6th instars (pers obs); and in this regard they are similar to the comparative instars of E. gillettii (Williams et al. 1984). The 3rd instar is brown bodied and there are two morphs on Islay – one appears pale overall (brown above and pale yellow–brown below with pale spines and a black head) and the other wholly dark (dark brown, black spines, black head). Subsequent instars are predominantly black with black spines, turning purple-brown close to ecdysis, with increasing amounts of white or silver markings on the dorsal surface. Fresh moults have pale heads and spines for a while.

Caterpillar development

The growth of caterpillars in webs was measured in the autumns of 2014–2016 and springs 2014 and 2015 to determine the standard weights and durations of each instar and observe any unusual patterns of development. In autumn, this encompassed the period from which caterpillars became prominent in the above-ground web-building stages, usually as late 2nd instars, until hibernation as 4th instars (mid-August to early October). In spring, caterpillars were sampled throughout development after emergence from hibernation as 4th instars until pupation (mid-February to early May).

Autumn sampling

Samples of caterpillars were taken by brushing a number from a web, usually 15–30, with a soft paintbrush into a small plastic pot. These were transferred to a 30 ml waxed paper cup that had been tared on a portable field balance (Ohaus Scout Pro, accuracy ± 3 mg), and the total mass and number of caterpillars was recorded. The instar of caterpillars in samples was also recorded and different instars were weighed separately. The largest caterpillars in each sample, and any that had just moulted, were weighed individually. Larvae were returned to their webs of origin. Up to 20 webs were sampled randomly on every site visit, according to availability, and sites were visited every 5–7 days.

Overall, 20,898 caterpillars from 1069 webs at 25 sites were sampled during autumns 2014–2016. Of particular interest were the weights and behaviours of caterpillars preparing for hibernation and those in hibernation. Searches for hibernacula were undertaken in autumns 2014 and 2015 and caterpillars within them were sampled and weighed.

Spring sampling

Sampling during the spring web phase, when 15–20 caterpillars per web were weighed, was conducted in the same manner as autumn, but the approximate lengths of unrolled caterpillars were also recorded. Of particular interest in the spring was the time taken by 4th instars to develop to moult after emergence from hibernation, and the weights at which this moult occurred as well those of 5th instars immediately after it. During the moult, 5th instars remaining in webs and in 1 m quadrats around them (up to a maximum of 20 caterpillars), were collected, counted, weighed together and the maximum recorded. New moults were weighed individually. As these dispersed subsequently, caterpillars were collected in random 1 m quadrats and weighed individually (up to a maximum of 20 caterpillars per quadrat). Overall, 9621 4th instars from 767 webs, 3641 5th instars and 3626 6th instars were weighed from 35 sites in springs 2014–2015.

Advanced caterpillars

Caterpillars that were unusually well-developed at the start of spring, being 5th instars during the early growth of 4th instars soon after emergence from hibernation, are referred to hereon as ‘advanced’ caterpillars. Searches for these advanced caterpillars were conducted at study sites at the beginning of each spring (mid-February until the moult of 4th instars), especially the general locations of the previous spring’s webs. Any observed were collected, weighed individually, photographed and the instar recorded. In spring 2014, three groups of study sites were selected based on the status of E. aurinia in previous seasons. Group A comprised the only two sites (in two 1 km squares separated by 18 km) that were occupied by E. aurinia in autumn 2012 and spring 2013 but at which no webs could be found in autumn 2013 (i.e. caterpillars should be absent in spring 2014); Group B were locations with webs in autumn 2013 but not in previous seasons (hence only webs of young caterpillars should be present, n = 15); and Group C were sites that had been occupied in all previous seasons (n = 18). Weights of caterpillars were recorded every 6–7 days from 22nd March 2014 until the spring moult of 4th instars in mid-April in what was a late season.

There were no Group A sites in spring 2015 owing to an abundance of E. aurinia in autumn 2014. The spring development of larvae in marked webs and any advanced caterpillars that were located during site searches was assessed at 11 sites (in 11 1 km squares). Three to five visits were made to each site from 8th March until the moult of 4th instars in early April. In 2016, the study focused on captive rearing and manipulations of caterpillar distribution in the field, but observations of the occurrence of advanced caterpillars at study sites continued on an informal basis.

Captive rearing

The development of 4th instars and advanced caterpillars collected in the field was observed in captivity in order to: (a) observe their comparative development; and (b) to investigate whether the latter were parasitised (braconid wasps Cotesia sp are common parasitoids of the larval stages of E. aurinia, Porter 1983), malformed or diseased and were unable to complete development. All consequent adults were released to their site of origin.

In the early spring of 2016, larvae were collected from sites soon after they emerged from hibernation and reared to the cessation of activity (pupation, dormancy or parasitism). Five entire webs of 4th instar caterpillars that had been marked the previous autumn, containing 160 larvae, were collected from two sites, and 60 advanced caterpillars were collected during searches of 12 sites from 15th February to 14th March 2016. Caterpillars were reared outdoors in containers with net covers (bath tubs and sheep-lick tubs containing sods of habitat supplemented with foodplant collected from study sites). The vegetation of containers was searched for caterpillars after the cessation of activity.

In 2015, ten advanced caterpillars from five sites were collected in early spring (10th-23rd March 2015) along with a small sample of 4th instar caterpillars (n = 15) from two sites. In the late spring (28th April to 2nd May) a sample of late, underdeveloped 5th instars (n = 15) was collected from the same sites. These were assumed to be parasitised as most caterpillars had pupated.

Manipulation experiments

Field experiments were conducted at three sites in 2015–2016 with the objective of confirming a prolonged life–cycle in E. aurinia. These rendered areas devoid of the generation of caterpillars that resulted from the flight period of 2015 and observed whether advanced caterpillars occurred in these in spring 2016. At one site (Cachlaidh Mhor), two plots that comprised 10 m squares 60 m apart were established in spring 2015 around concentrations of caterpillars. Searches for webs of E. aurinia were undertaken in autumn 2015 inside and around the plots during repeated visits – no webs occurred within the plots and one web that was found between them was moved.

Four exclusion plots were established at two RSPB reserves (Creag Mhor and the Mull of Oa) in June 2015 just before the flight period of E. aurinia. Plots were located in areas in which webs had occurred in autumn 2014 and spring 2015. Fine netting, 10 m by 8 m, was laid on each plot to exclude butterflies during the flight period (Fig. 3, butterflies that eclosed under the netting were released). Searches for webs were made at each plot throughout autumn 2015. No webs were located inside them, and webs within 5 m of their perimeter were moved to two holding areas at least 50 m from the exclusion plots to reduce the likelihood of caterpillar ingress. Wire fencing was erected around each exclusion plot during the winter of 2015/2016 to exclude livestock (cattle graze the sites in November) and then barriers were erected around the perimeters of the plots in early February to limit the ingress of caterpillars in spring. These were continuous lengths of fine mesh – 50% windbreak netting composed of a triangular mesh with a maximum aperture of ca 1.5 mm, folded in half and attached to and held upright to a height of 1 m by electric fence posts and driven into the ground with tent pegs to form a seal (Fig. 3). The integrity of the netting was checked periodically.

Fig. 3
figure 3

One of the exclusion plots at Creag Mhor RSPB reserve, Loch Gruinart, Isle of Islay, that was used to manipulate the distribution of caterpillars, showing the netting used to exclude butterflies during the flight period (left) and the exclosure erected the following year to limit the ingress of caterpillars (right)

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All six plots were monitored during the spring from 15th February 2016. Caterpillars occurring within them were weighed and their development was compared with caterpillars of known age in re-located webs in the holding areas and in webs that had been marked on site during autumn 2015. All caterpillars were weighed individually. Thereafter, the manipulation plots were re-netted during the flight period of 2016 and caterpillar barriers re-erected in spring 2017, following the same procedures as 2015–2016.

Results

Key weights of instars

Each instar roughly doubles its length and quadruples its weight (Fig. 4). The moult of 2nd instars was usually over by late August although in 2015 webs of these were still evident in mid-September. Second instars moulted at 2-3 mg and 3rd instars took 4–6 weeks to reach moulting weights of 10-12 mg. Fresh 4th instars still within moult webs with 3rd instars weighed on average 6.4 mg (n = 1412). Once moulted, 4th instars did not feed and they constructed another web high in the vegetation, usually suspended by grass stalks, in which they prepared for hibernation. Caterpillars in these webs remained at 6.4 mg (n = 3763). Only 32 pre-hibernation 4th instars from the entire sample (5174) weighed > 10 mg, the largest individual being 15 mg.

Fig. 4
figure 4

The relationship between E. aurinia instar length and weight: 4th (white circles, n = 248); 5th (black squares, n = 83); and 6th (white triangles, n = 122). Larvae hibernate at the beginning of the 4th instar

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Caterpillars spent up to 10 days or so in the pre-hibernation web, but then constructed a tight spherical web deep in the vegetation beneath this in which they hibernated. Caterpillars in hibernation in October weighed on average 6.2 mg (n = 852 from 51 hibernacula) and only one hibernating caterpillar was found that weighed more than 10 mg (12 mg). All bar two were 4th instars (two 3rd instars were found in hibernation webs), and no solitary caterpillars occurred in the autumn at any site.

Some 4th instars emerged from hibernation in February and constructed webs on which to bask but did not feed immediately. Most had appeared by mid-March in each year studied when they weighed 4-8 mg. Moulting weights of around 25 mg were attained after about four weeks, including a few days in which they prepared for and underwent moult (so moult was usually late March to mid-April). Fresh 5th instars weighed 15-23 mg (mean 19.5 ± 0.4 mg, n = 35), took about two weeks to reach moulting weights of 80-100 mg, and new 6th instars weighed 76.3 ± 2.6 mg (n = 20). The 6th instar also took about a fortnight to reach pupation, attaining weights in excess of 300 mg (Fig. 4).

Comparative spring development of caterpillars

2014

No webs of 4th instars occurred at the two Group A sites but two solitary 5th instars were found at one of the sites on 25th March 2014, weighing 42 and 22 mg, well before 4th instars moulted elsewhere (Fig. 5). Further 5th instars occurred at this site before 15th April weighing 57-133 mg. No caterpillars were found at the other Group A site until 10th April when five 5th instars were found, weighing 51.0 ± 5.2 mg.

Fig. 5
figure 5

Spring 2014: the mean weights per day ± SE of advanced caterpillars at Group A locations (black squares) compared with the growth of 4th instars (grey) and subsequent 5th instars (open squares) in and around webs at Group B locations

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No advanced 5th instars and only 4th instars in webs were recorded at Group B sites in March. The first moult of 4th instars at these was recorded on 5th April, but new 5th instars were not frequent until the 12th April. The mean weights of 5th instars at these sites did not exceed 60 mg before the 15th April (Fig. 5). Both 4th instars and advanced caterpillars occurred at four of the 14 Group C sites (and only 4th instars at the remainder) – nine advanced caterpillars weighing 38-126 mg occurred before webs of 4th instars had started to moult at these sites.

2015

Advanced caterpillars occurred at five of the 11 sites. The earliest weighed 30 mg on 10th March while 4th instars were 5.5 mg (n = 53) and emerging from hibernation (Fig. 6). Thirteen were recorded on 13th March (mean 29.0 ± 2.0 mg), when 4th instars were 6.6 mg (n = 299), and 15 were found on 16th March (mean 38.3 ± 1.5 mg) when 4th instars were 6.3 mg (n = 77). The first moult of 4th instars at these sites was recorded on the 27th March, continuing to the 12th April. The development of caterpillars at the six sites without advanced caterpillars was slightly faster and more synchronised, reaching moult on the 25th March and finishing by 3rd April (Fig. 6). The largest 4th instar recorded at all sites in the first two weeks of March was 15 mg (n = 814).

Fig. 6
figure 6

Spring 2015: the mean weights ± SE of co-existing advanced caterpillars (black, diamonds show samples with final instars) and 4th instars (open circles) and subsequent 5th instars (grey circles) at five sites; and 4th instars (open triangles) and subsequent 5th instars (grey triangles) at six sites without advanced caterpillars

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2016

The first advanced caterpillars, primarily solitary but also in small groups of 2–6, were recorded on 15th February and they occurred at five sites by the end of the month. Their mean February weight was 33.6 ± 2.1 mg (n = 14) and between 7 and 14th March they weighed 50.6 ± 2.7 mg (n = 24). Only three webs of 4th instars were located in February and caterpillars in these webs weighed 6.9 ± 0.3 mg (n = 23, maximum 10 mg). Most did not emerge from hibernation until after 3rd March and the moult started on 27th March.

Captive rearing

Viability of advanced caterpillars

Two of 70 collected advanced caterpillars were parasitised (one of 10 in 2015 and one of 60 in 2016). Eight larvae of C. bignellii emerged on 7th April 2015 from a 5th instar weighing 72 mg and six C. bignellii larvae emerged on 15th April 2016 from another of 80 mg. The remaining advanced caterpillars developed and pupated.

Development of caterpillars

Advanced caterpillars collected in February 2016 weighed 33.6 ± 2.1 mg (n = 14). They did not feed initially and developed slowly in the first half of March, thereafter accelerating to start moult to the final instar at the end of the month (Fig. 7). The mean weights of 4th instars in webs were 5.7–11.0 mg on collection in 2016 and did not exceed 20 mg until 4th April. These started moulting on 9th April (Fig. 7).

Fig. 7
figure 7

Spring 2016: the mean weights ± SE of advanced caterpillars (black, diamonds show samples containing final instars) and 4th instars (white circles) and subsequent 5th instars (grey circles) in captivity

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Five 4th instars developed slowly in 2016 and became inactive as 5th instars in sheaths of loose silk attached to vegetation during May – four in a tight group and one individually. These weighed 55.0 ± 6.7 mg and 67 mg respectively. All moulted during this process and retained the morphology of the 5th instar. Another caterpillar became inactive as a 4th instar – this was placed in a small pot where it spun a web of silk and weighed 14 mg on 27th May. No other 4th instars were found to remain in rearing containers after activity ceased. All inactive caterpillars were lost to mould later in 2016.

In spring 2015, two caterpillars collected as 4th instars developed slowly and stopped feeding as 5th instars in late spring weighing 33 mg and 42 mg, spun webs individually and became inactive – the latter weighed 33 mg on 9th August 2015. Another remained as a 4th instar and became inactive on 16th April weighing 16 mg but died soon after. Underdeveloped 5th instars collected in late-spring 2015 weighed 46.0 ± 3.0 mg (n = 15). Six moulted to the 6th instar between 3rd and 15th May – four pupated subsequently and two were parasitised. All wasps from these caterpillars were C. melitaearum. The remaining 5th instars became inactive and weighed 24-48 mg (mean 39.7 ± 2.7 mg) on 16th May. By 25th May there were two groups, of four and five, in balls of silk. Individuals from these webs occasionally became active between 10th July and 22nd September and left the web briefly. These weighed 22-33 mg (mean 28.7 ± 1.6 mg, n = 6). Webs and caterpillars were extant in October 2015 but were eventually lost to mould.

Manipulated field plots

Cachlaidh Mhor

Under-developed 5th instars were common at Cachlaidh Mhor when the plots were established in late-spring 2015 at a time that 6th instars are usually well-developed or pupating – 73.8% were still 5th instars on 28th April (n = 143) and 52.9% on 6th May (n = 86). These weighed 64.6 ± 2.2 mg (n = 74) and 68.3 ± 2.6 mg (n = 45) respectively. They were still evident on 13th May (62.1 ± 3.8 mg, n = 18) by which time they had gathered in small groups.

Six small groups of these 5th instars (ca. 3–6 individuals) occurred within the two plots at Cachlaidh Mhor and these constructed webs. All were extant on 12th June 2015 and two webs were opened – these contained 4–6 caterpillars that had moulted as exuviae were present but they retained the morphology of the 5th instar. By 21st June, caterpillars had moved and constructed spherical webs deep within the vegetation and all six groups were extant on 4th August 2015 and their locations marked.

No webs of 4th instars occurred in the plots in spring 2016 (when only two webs could be found on the whole site). The first advanced caterpillars, 5th instars weighing 28, 38 and 39 mg, emerged on 15th February 2016 and three more groups were by markers in plots on 23rd February. Thereafter, solitary 5th instars were frequent at this site – 83 advanced caterpillars were recorded on a 297 m transect at this site on 14th March (and only two webs of small 4th instars).

RSPB reserves

No webs of 4th instars occurred in any of the plots in spring 2016. Two advanced caterpillars occurred within the plot at the Mull of Oa on 23rd February 2016 (5th instars of 29 mg and 32 mg), one within a plot at Creag Mhor on 25th February and another in the same plot on 27th February (32 mg). Thereafter, one to three advanced caterpillars, always solitary 5th instars, occurred in each RSPB plot bar one during searches in March (a final instar was found subsequently in the vacant plot). No advanced caterpillars occurred within the plots in spring 2017.

Caterpillar development

The growth of advanced caterpillars in plots was initially slow as they still weighed 36.7 ± 2.9 mg (n = 20) on 14th March 2016 but they were 84.9 ± 4.4 mg (n = 20) on the 5th April and had started the moult to the final instar on 30th March (Fig. 8). Caterpillars in translocated and marked webs showing in February at the experimental sites weighed 5.3 ± 0.3 mg (max 7 mg) and 13.6 ± 0.5 mg by the end of March (max 19 mg). The earliest moult to the 5th instar occurred on 7th April.

Fig. 8
figure 8

Spring 2016: the mean weights ± SE of advanced caterpillars (black, diamonds show samples containing final instars) within manipulated plots; and 4th instars (white circles) and subsequent 5th instars (grey circles) in and around re-located and marked webs at the same sites

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Discussion

The longer life-cycle of E. aurinia

The weight of advanced caterpillars of E. aurinia at the beginning of spring was consistent in all years and roughly five times that of caterpillars known to be in their first spring. The subsequent comparative development of larvae was also the same in all springs and situations studied – it took more than a month for caterpillars in their first spring to develop to the weight and instar at which advanced caterpillars occurred. There was no indication that some larvae might develop rapidly and are unusually large before hibernation in autumn, let alone reach the 5th instar, and all caterpillars entered hibernation as fresh 4th instars (bar two 3rd instars). These data alone indicate that advanced caterpillars must be older, and the subsequent observations that some caterpillars develop slowly and become dormant in late spring at weights that correspond with the weights of advanced caterpillars in early spring, and that advanced caterpillars occur in areas rendered devoid of young caterpillars, confirm that E. aurinia is able to add a year to the life-cycle in Scotland.

Those species within the Melitaeini tribe of butterflies (that includes E. aurinia) that can extend their life-cycle do so by entering diapause after a period of post-hibernation growth (Williams et al. 1984; Luckens 1985; Wahlberg 1998; Eliasson and Shaw 2003; Pratt and Emmel 2010). Most of these enter diapause in the 5th instar e.g. E. maturna (Wahlberg 1998), E. gillettii (Williams et al. 1984) and E. intermedia (Luckens 1985). Scottish E. aurinia are similar therefore, but the additional moult of E. aurinia, increasing the number of instars in the life-cycle to seven, seems not to have been recorded before. An extra instar is a feature of insects that extend their life-cycle (Esperk et al. 2007), and this extra instar of E. aurinia, added when caterpillars are preparing for diapause, is identical to the 5th instar.

In Sweden, most larvae of E. aurinia that add an extra year do so as aggregations of 4th instars (after only a brief appearance in their first spring) and only those that attempt a third year do so as 5th instars (Eliasson and Shaw 2003). The possibility that advanced caterpillars on Islay were in their third year cannot be discounted, as webs of 4th instars were found sometimes in early spring where they had not been recorded the previous autumn. Two 4th instars also appeared to enter diapause in spring in captivity, although both died during the summer. However, the caterpillars that entered diapause as 5th instars in captivity in spring 2016 originated from webs marked the previous autumn, so must have been in their first year (although mixing of 4th instars in the early spring cannot be discounted). Furthermore, no 4th instars occurred during spring in any of the field plots that were manipulated the previous year and none remained in containers after captive rearing, despite searches. The current weight of evidence suggests that only the 5th instar of E. aurinia (or technically the 6th as they moult before diapause) extends the life-cycle in Scotland. Whether this can add further years is unknown, although no advanced caterpillars reared in captivity re-entered diapause and none were found in field plots after a further year’s manipulation. Despite these observations, the possibility remains that the life-cycle of E. aurinia may be longer in the UK than recorded in this study given the relatively small numbers of larvae observed in experimental plots.

Longer-lived caterpillars are very characteristic in the early spring on Islay. They can be located from mid-February at characteristic weights of 25-40 mg (when spring 4th instars, if they have emerged, weigh 4-8 mg), having entered diapause the previous spring at weights of 40-60 mg. They are shrivelled on emergence from weight loss, through dehydration presumably and the suspension of metabolism. This lends them a very black and especially spiky appearance, caused by the compression of body segments and spines, with a prominent head capsule, which in the 5th instar is already relatively large at 1.5 mm. They are usually solitary and difficult to find, as they do not feed immediately but bask for long periods and are otherwise cryptic, presumably whilst they regain function. One advanced caterpillar was not seen, and nor could it be found, after placing it in a small tub of vegetation on 22nd February 2016 when it weighed 27 mg, until 20th March when it weighed 39 mg – but it pupated at 257 mg on the 23rd April. They reach the final instar (technically the 7th) at about the same time as 4th instars are moulting to 5th instars, and this produces a characteristic diversity of caterpillar size and instars at sites with a prolonged life-cycle. The size and development of caterpillars at sites at which no advanced caterpillars were recorded was more uniform and synchronised.

Conservation

The longer life-cycle of E. aurinia may permit some individuals to by-pass events which may cause the failure of the remaining population and enhance the persistence potential of the species. The extra instar achieved by caterpillars on a longer life-cycle may also be significant, as insect larvae that add instars while growing over extended periods usually attain greater weights, which in turn are reflected in adult size, mobility and fecundity (Berger et al. 2006; Esperk et al. 2007; Etilé and Despland 2008; Saastamoinen et al. 2013).

The possibility that populations of E. aurinia in the UK may have a longer-life-cycle has not been considered previously, perhaps as insects with biennial or multiennial life-cycles tend to show regular peaks of abundance or synchronised emergence (Várkonyi et al. 2002; Kleckova et al. 2015). Factors such as weather and the impacts of parasitoids, as well as the effects of habitat quality and management, have been implicated in the dramatic fluctuations that are characteristic of the species (Porter 1983; Lei and Hanski 1998; Asher et al. 2001; Bulman et al. 2007). It is conceivable that the longer life-cycle of E. aurinia may contribute to volatility if there are years in which a proportion, if not all, of the population enters diapause instead of developing to pupation. This will render it difficult to observe in some years, and indeed the species is known for its propensity to escape detection for long periods (Ford 1930; Porter 1990; Ravenscroft 2019).

The longer life-cycle means that caterpillars may be present at sites despite an absence of the species in summer and autumn. This may complicate the understanding of the structure and function of E. aurinia populations gained from studies of population turnover (Schtickzelle et al. 2005; Bulman et al. 2007). This is especially so of studies that assessed extinction and colonisation (or vice versa) in consecutive years, some of which reported the occurrence of adults following an absence of larvae in autumn (Wahlberg et al. 2002; Anthes et al. 2003; Hula et al. 2004). It has implications for standard monitoring and other assessments of the species too, which are carried out on young caterpillars in their first autumn (Lewis and Hurford 1997; Asher et al. 2001). However, its occurrence elsewhere in the UK is unknown currently – short searches of mine on mainland Scotland have failed to find it, as did a short study of Irish populations in 2017. But Islay is a relatively flat island and its oceanic climate is mild (and wet and windy). The periods of growth of E. aurinia caterpillars on the island seem little different from those reported elsewhere and there is no reason to believe that the longer life-cycle is not more widespread. Few insights are possible from the literature as caterpillar ecology in the spring has been little studied. An exception is a study by Porter (1983) of the relationship between E. aurinia and its parasitoid C. bignellii at a site in Oxfordshire, England at which E. aurinia is now extinct. Porter recorded unusual occurrences of C. bignellii emerging early in small batches from 5th instars in the spring of one year. This bears a striking resemblance to the emergence of C. bignellii from caterpillars after a year in diapause on Islay, and is a tantalising suggestion that a longer life-cycle may exist, or have existed, in southern England.

The extended life-cycle did not occur (or was not found) at all sites on Islay in this study. This suggests that either the protracted diapause of caterpillars occurs only at sites with particular conditions (or it is just more successful at these) or that it may be more widespread only in particular years. The mortality of E. aurinia is greatest during hibernation (Liu et al. 2006), so specific qualities of habitat are probably important to caterpillars attempting an even longer-term diapause (e.g. E. editha, Pratt and Emmel 2010). Appropriate humidity is thought to be critical to Euphydryas caterpillars in diapause (Eliasson and Shaw 2003). The species does occupy a wide range of habitat conditions in Scotland, some of which do not conform to recognised parameters of quality. The vegetation of sites at which longer-lived caterpillars occurred on Islay was usually relatively tall, dense and damp, often with a deep ground layer, such as bryophytes and litter, and with relatively little foodplant S. pratensis as a consequence. An abundance of the foodplant is regarded as the principle feature of high quality habitat (Fowles and Smith 2006; Betzholtz et al. 2007; Smee et al. 2011; Botham et al. 2011). If it does transpire that the prolonged life-cycle is only successful under certain conditions and that the features of these habitats differ from recognised qualities, then the relevance of the prolonged life-cycle to the long-term persistence of E. aurinia will need to be determined before implications for conservation management can be understood. Longer life-cycles are not always significant in the long-term to insect populations with the capacity for them (Vila and Björklund 2004) and in any case the application of current knowledge of conservation management for E. aurinia is generally successful (Porter and Ellis 2011).