Down the up staircase: Equatorward march of a cold‐water ascidian and broader implications for invasion ecology

While warming temperatures are expected to facilitate the poleward movement of species previously restricted to more equatorial waters, the arrival and persistence of cold‐water species in more equatorward waters are relatively unprecedented. The native north‐east Pacific ascidian Corella inflata Huntsman, 1912, has spread southward and invaded new regions along the North American Pacific coast, a rare example of a marine species moving towards the equator. Here, we document C. inflata's equatorward movement and potential impact, assess several hypotheses for its spread and consider implications for invasion ecology.


| INTRODUC TI ON
The poleward range expansions of numerous lower-latitude terrestrial, freshwater and marine species during the past half-century are now well documented (Bates et al., 2014;Canning-Clode & Carlton, 2017;Chen, Hill, Ohlemüller, Roy, & Thomas, 2011;Sunday et al., 2015). These movements are likely underestimated due to the same challenges that limit the recognition of species invasions, including limited search effort and constraints of taxonomic and biogeographic knowledge (Carlton, 2009). With rare exceptions, these northbound in the Northern Hemisphere and southbound in the Southern Hemisphere range expansions are correlated with warming land and water temperatures. Exceptions may include cases in which new habitat becomes available, suggesting that species' geographic boundaries are in some cases limited by substrate and not temperature. One such example, albeit not a poleward extension, may be the historical southern expansion of the intertidal barnacle Semibalanus balanoides, which in the mid-1900s extended south along the North American mid-Atlantic seaboard after the construction of rock groins on long stretches of sandy shores (Carlton, Newman, & Pitombo, 2011); this barnacle is now retreating north with warming temperatures (Jones, Southward, & Wethey, 2012).
Numerous non-native animals and plants may spread both north and south along a coastline after introduction to a new region, a phenomenon typically correlated with a given species' thermal tolerance limits (Epifanio, 2013;Grosholz, 2010;Reid, 1996).
In a steadily warming world, the expansion of a higher-latitude marine cold-water species into warmer-temperate waters is unexpected, given that such species are presumably evolutionarily restricted to lower temperature waters for reproduction, settlement, growth and survival. We consider here the curious case of the native north-east Pacific Ocean solitary ascidian (Corella inflata Huntsman, 1912 [Ascidiacea, Phlebobranchia, Corellidae]), whose southern limit was long recognized as Puget Sound, Washington. Commencing in the 2000s, C. inflata began appearing in Oregon and California ports and harbors that had long been under extensive biological scrutiny.
Corella inflata ( Figure 1) is a conspicuous, transparent, cuboid species reaching 5 cm in length. It was first described "from between tides to about 10 fathoms" (18.3 m) from Departure Bay, Nanaimo, British Columbia (Huntsman, 1912). Van Name (1945) mistakenly considered C. inflata to be a synonym of the older-named Corella willmeriana Herdman, 1898, a larger species reaching deeper waters, believing the former to be smaller specimens of the latter. Based upon morphological and reproductive evidence, Lambert, Lambert, and Abbott (1981) resurrected C. inflata;all literature between 1945 and 1981 thus confused the two species, referring to C. inflata as C.
willmeriana. Lambert et al. (1981) reported the range of C. inflata to be from the northern tip of Vancouver Island (Hope Island, Queen Charlotte Strait), British Columbia to Puget Sound and the usual range of C. willmeriana to be nearly identical, from southernmost (south-east) Alaska (Loring) to Puget Sound. A few confirmed specimens of C. willmeriana were taken 50 years ago in Monterey Bay, California, at depths of 30 and 60 m (Lambert et al., 1981), and were the basis of the comment in Lambert, Lambert, and Lambert (1995) that C. inflata ranges "to central California." This unusual record is presumably the result of a rare transport event of the species from the Pacific Northwest to Monterey Bay, and is unrelated in time or space to the 21st-century phenomena that we describe here.
Recently, since the 1990s, C. inflata has also been found far to the north in south-central Alaska. It was discovered in Prince William Sound in 1998Sound in -1999 where it was common in some marina float fouling communities (Hines & Ruiz, 2000a;Lambert & Sanamyan, 2001;Ruiz et al., 2006), and then further west in Kachemak Bay in 2000 (Hines & Ruiz, 2000b). Subsequent reports filled in the range in south-eastern Alaska, including Sitka Sound (rare, 2007: Pirtle, Ibarra, & Eckert, 2012, Bartlett Cove, Glacier Bay (common, 2010: M. Noble andL. McCann, personal communications, 2012) and Ketchikan (abundant, 2009: Davis, 2010. The report by Bluhm, Iken, Hardy, Sirenko, and Holladay (2009) Rosenthal, Lees, and Rosenthal (1977) reported "Corella willmeriana" earlier from Zaikof Bay, Prince William Sound, based upon 1975Sound, based upon -1976 collections, but without descriptive or other details; as their report pre-dates the 1981 revision of Corella taxonomy, the record (if correct) could represent either C. inflata or C. willmeriana. Lambert et al. (1981) remarked that "very extensive collections carried out (north of southernmost Alaska) and the Canadian Arctic have all failed to discover a single specimen of Corella." We suggest F I G U R E 1 Corella inflata fouling a submerged bucket in Coos Bay, Oregon, October 2010 (photograph by Gretchen Lambert) that the above northern records of C. inflata and C. willmeriana likely reflect climate-induced expansions, as would be expected in the latter half of the 20th century, possibly as early as the 1970s, but more certainly during and since the 1990s.
Here, we document the anomalous southward range expansion of C. inflata (hereafter, Corella). We further examine several hypotheses for this expansion, and report upon possible impacts, including changes observed in community composition and structure in San Francisco Bay after Corella appeared.

| Distribution of C. inflata south of Puget Sound
We assessed the known distribution and southward movement of C. inflata along the west coast of North America using three data sources. We compiled records of C. inflata occurrence from the literature and from correspondence with workers along the coast from Alaska to southern California. We conducted opportunistic, informal searches of floats and pier pilings for Corella, following the discovery Standardized surveys were done at least once in each bay, and repeated multiple times in several bays, including San Francisco Bay, which was surveyed for 18 consecutive years.
Initially, Corella were identified morphologically from multiple sites along the west coast of North America (Lambert 1981;G. Lambert, personal communication), and the morphology of spec-

| Standardized fouling community survey methods
We compiled results from standardized three-month panel studies of sessile invertebrate community development at 6 to 10 sites in each of 28 bays along the Pacific coast of North America, including 17 bays south of Corella's original southern range boundary at Puget Sound (see results for locations and years). All surveys were conducted during the summer months (June-September), coinciding with the season of high recruitment, to assess community composition (deRivera et al., 2005; Simkanin et al., 2016;G. M. Ruiz and A. L. Chang, unpublished data). For these standardized surveys, 14 cm × 14 cm × 0.5 cm square, grey, polyvinyl chloride (PVC) panels were used as passive recruitment collectors. Panels were distributed throughout each site using a stratified-random design. Each panel was lightly sanded, attached to a brick for weight and suspended panel side-down from a rope tied to a dock or buoy. The panels were examined for the presence of benthic marine invertebrates, following standard methods described below for San Francisco Bay. Here, we report only on Corella presence in these surveys across bays.
As part of a long-term study in San Francisco Bay, we conducted systematic panel-based surveys of epifaunal community composition over an 18-year period (2000-2017) to detect seasonal and interannual changes in community composition (Chang, Brown, Crooks, & Ruiz, 2018). To study sessile invertebrate community de- In San Francisco Bay, panels were left in place for 12-14 weeks (summer and quarterly panels) to record invertebrate settlement on the downward-facing side, then retrieved and replaced with new, blank panels at the same location. Community development as measured here thus encompasses both settlement and post-settlement processes, including mortality, which occurred during the entire deployment period. After retrieval, panels were analysed for species composition, per cent cover and biovolume. To estimate per cent cover of dominant taxa, a grid of 50-100 points was placed over each panel and the taxon attached to the panel at each point (i.e., the "primary" cover organism) was identified to the lowest possible taxonomic level using a dissecting microscope. If other organisms were growing on top of the primary cover organism at a point, these "secondary cover" organisms were also identified and recorded. Total per cent cover was the sum of primary cover and secondary cover and could thus exceed

| Environmental measurements in San Francisco Bay
We monitored temperature at 1 m depth at each site, while survey panels were deployed using a combination of Hobo loggers (Onset

| Historical temperature data
To examine hypotheses about relationships of C. inflata population dynamics to longer-term temperature changes, we examined records of water temperature data from locations near where we found

| Statistical methods
In San Francisco Bay, we compared community composition at the two sites (San Francisco Marina and Sausalito Marine Harbor) where Corella was eventually observed and for which we had extensive data before and after Corella's appearance. We used oneway PERMANOVA routines performed on Bray-Curtis dissimilarity matrices of square root-transformed per cent cover data of the taxa on each panel at every available time point from summer surveys at these sites. Corella itself was omitted from the data to examine the impact on the other species in the community; there also was no secondary cover on Corella. Square root transformation was used to reduce the effects of extremely abundant taxa while simultaneously emphasizing the effects of rare taxa. The matrices were visualized using non-metric multidimensional scaling (nMDS). We then used SIMPER, a similarity percentage procedure, on Bray-Curtis dissimilarity matrices of square root-transformed per cent cover data to ascertain which species were most responsible for community differences after the Corella invasion. Separate analyses were performed for each site. Shannon diversity was calculated for each panel.
Community composition on San Francisco Bay survey panels was also compared to environmental parameters using PERMANOVA.
We tested for the effect of environmental conditions during community development by using the mean water temperature and salinity during the panel deployment as predictor variables. Because the previous winter's salinity levels have also been shown to significantly impact San Francisco Bay fouling communities (Chang et al., 2018), we also used the mean February-to-May Net Delta Outflow as predictor variables to explain variation in Corella presence and abundance on summer and quarterly panels at each site.
We used least-squares linear regression to evaluate the sign and

| Distribution and current status of C. inflata south of Puget Sound
Corella inflata was first discovered south of Puget Sound in the fall of 2004 in Coos Bay, Oregon (Table 1;
Numerous changes to existing communities were observed fol-  Figure 6). Using SIMPER to examine the species other than Corella that were most responsible for community differences after the invasion, we found that the abundances of a wide range of taxa changed, with more pronounced differences at Sausalito Marine Harbor than at San Francisco Marina (Table 5).

| Temperature trends in Friday Harbor, Coos Bay and San Francisco Bay
During the time period tested (1993-2016), annual mean tempera-  Corella inflata retains its embryos in an enlarged brood chamber (Lambert et al., 1995)

| Distributional ecology and community impact in San Francisco Bay
Starting in June 2008, C. inflata was detected consistently, but at varying abundances, at two sites near the mouth of San Francisco Bay. Given the broad geographic scope and frequent repetition of our surveys within the estuary, it seems likely that Corella's distribution within the Bay remained limited to the general vicinity of areas we first found it. Larvae were observed in the brood chambers of adults each year on quarterly, summer and long-term panels, with new settlers appearing in spring and early summer (Figure 4).

Corella's incidence and abundance after detection at San Francisco
Marina and Sausalito Marine Harbor were strongly correlated with environmental conditions, with greater abundance in slightly lower (but not below 20 ppt) salinity levels. The significantly greater abundance recorded during years with intermediate freshwater flow entering San Francisco Bay suggests a possible preference for slightly lower salinity conditions. The abundance of potential competitors was not significantly greater during drier years (higher salinity conditions), suggesting TA B L E 1 North-to-south records, south of Puget Sound, of the ascidian Corella inflata compiled from literature surveys, correspondence, informal surveys and standardized panel surveys (see Tables 2 and 3 Figure 6). We observed monolayers of Corella on quarterly, summer and long-term survey panels. Young (1988) found that such monolayers could form because Corella consume larvae of other species (as well as conspecifics), but have themselves evolved mechanisms to avoid settling on conspecifics. As year than these other species-as would be expected from its Pacific Northwest origins-thus fitting into a habitat facies that was poorly occupied by other solitary ascidians.

| A Corella Novella: why a cold-water species would colonize warmer-temperate latitudes
We presume that C. inflata has long been transported via ships out of the Pacific Northwest, but was not detected elsewhere along the Pacific coast (or the world) until the beginning of the 21st century, despite possessing a number of attributes associated with Note: The total number of panels examined from each bay in a given year is specified in each cell. Shading indicates that C. inflata was detected. See text for description of replication and study design.  colonizing species. Corella is a common-to-abundant fouling organism on floats, docks and piers (Cordell, Levy, & Toft, 2013;Lambert, 1968;Lambert et al., 1981;records herein), making it likely to foul vessels. Well before it was resurrected as a distinct species, Corella was noted as the "float" or "shallow-water" form of C. willmeriana (Lambert et al., 1981). As noted above, Corella's brooding life history includes ovoviviparous reproduction in which tadpole larvae are released and settle gregariously (Lambert et al., 1995), both facilitating colonization and quickly leading to dense monocultures of adults ( Figure 1). Individuals can reach sexual maturity in 90-120 days at a body length of about 12 mm, with breeding occurring throughout the year (Lambert, 1968, as C. willmeriana;Jacobs & Sherrard, 2010 to detect such marine macrofauna over the past 60 years (Carlton et al., 1979;Ruiz, Fofonoff, Carlton, Wonham, & Hines, 2000;Ruiz, Fofonoff, Steves, Foss, & Shiba, 2011).

TA B L E 2 (Continued)
We examine four hypotheses for why Corella would be successful now, rather than historically, south of its apparent natural southern boundary.  Table 3 for years surveyed.
[Correction statement added on 29 May 2020 after first online publication: Alignment of numerical entries in Tables 2-5 have been amended in this version] Carlton (1992) noted that the invasion of San Francisco Bay by the Japanese semelid clam Theoralubrica occurred soon after this species increased in abundance in the Inland Sea of Japan. Clearly, larger populations of a given species in a donor region may provide greater opportunity for vector engagement. In the present case, we have found no data suggesting that Corella increased in abundance in the early 2000s or later years in the Pacific Northwest.
While Corella's abundance has likely fluctuated over time in northern waters, it did not appear until the early 2000s to the south. We thus find no evidence to support this hypothesis alone, although it could act in conjunction with a change in transport conditions, as described below.
2. Increased Turbidity in Bay Mouths South of Puget Sound Bingham and Reyns (1999) and Bingham and Reitzel (2000) demonstrated that C. inflata, with a thin, transparent tunic, is found primarily in areas where it is protected from exposure to direct sunlight. Decreased light penetration in shallow waters due to increased turbidity (due to a number of reasons, such as changes in suspended sediment loads, plankton or seston) could conceivably increase habitat availability for Corella. However, we found no data suggesting that the near-surface waters where Corella now occurs south of Puget Sound have become significantly more turbid. Moreover, under-float habitat, protected from ultraviolet radiation, has long been available in Oregon and California bays, but was not previously colonized by Corella. We thus also find no support for this hypothesis.

Decreased Bay-Mouth Temperatures in Oregon and California
All of the sites where this ascidian has been detected south of Puget Sound are in relatively shallow marina basins close to the mouths of each bay. Coastal water temperatures may cool locally due in part to more intense upwelling in Eastern Boundary Upwelling Systems (e.g., Bakun et al., 2015), which suggests that local near-  (Davidson et al., 2010;Floerl & Inglis, 2005). Recent studies have demonstrated significant vessel movement along the coast of California (Zabin et al., 2014). More broadly, Iacarella, Davidson, and Dunham (2019)  patterns, an understanding of the one or more processes that would serve to explain the equatorward movement of a cold-water affinity marine species remains elusive. We demonstrate that this ascidian has survived and reproduced over multiple years in waters well south of its previously known range, revealing a previously little understood breadth of its environmental tolerances and setting the stage for compelling experimental and genetic studies.

ACK N OWLED G EM ENTS
We are grateful to John Chapman, Sean Craig, Richard Emlet, Janet