Exactly 3ish American Galba

Editor’s Notes – This is the third in a four-part series prompted by the recent publication of our paper on genetic relationships in the worldwide lymnaeid genus Galba [1].  You will certainly find it helpful to review my essay of [7June21] before proceeding.  My essay of [22June21] is optional, depending upon how deep into this justifiably-obscure topic you actually want to dive.

This essay was subsequently published as: Dillon, R.T., Jr. (2023c)  Exactly Threeish American Galba.  Pp 137 – 149 in The Freshwater Gastropods of North America Volume 7, Collected in Turn One, and Other Essays.  FWGNA Project, Charleston, SC.

During the 2015 field season I did my best to keep the malacological pipeline between Charleston and Montpellier stuffed with samples of basommatophoran pulmonate snails, especially the crappy-little amphibious lymnaeids that we here in the USA often call “Fossaria,” but most of the rest of the world calls Galba [2].  In July I returned to the type locality of Thomas Say’s (1822) L. humilis (and L. modicella, oddly) on the bank of the Susquehanna River in the little town of Owego, NY [4].  I also visited Philadelphia to collect a topotypic sample of Lymnaea obrussa Say 1825, Cincinnati to sample topotypic Lymnaea parva Lea 1841, and Tennessee to sample topotypic Lymnaea exigua Lea 1841.  Most of the other specific nomina one occasionally sees in the literature for these little lymnaeids are more difficult to pin down geographically.  But I felt as though if I added samples from South Carolina, North Carolina, Virginia, and Michigan, genetic variation in North American populations of tricuspid fossarines should be adequately surveyed.

Topotypic L. (Galba) humilis [5]

I also sent a fresh sample of the bicuspid L. cubensis from Charleston to Montpellier, again harkening back to my [25June08] essay on our particularly-interesting local population.

And I very much enjoyed working with Dr. Pilar (“Pili”) Alda, the post-doc in the Hurtrez-Boussès lab spearheading the Galba survey.  In addition to her obvious scientific and technical skills, she turned out to be relentlessly cheery and outgoing, character traits that would serve her well dealing with the likes of me, to say nothing of her team of 18 other co-authors recruited from all over the world. According to my long-time friend Philippe Jarne, who often served as liaison, most of the field work was done by Jean-Pierre Pointier guided by colleagues “generally accompanying him for days to weeks in places where he would have a hard time going on his own, and also providing facilities (e.g. car, mules and lodging).”

Five years passed.  I knew that Pili’s analysis would involve our brand new multiplex PCR test [6], as well as more conventional morphological and DNA sequence data.  But regarding the myriad analytical details I was actually rather pleased to be out of the loop.  Finally, at long last, the paper appeared this spring in Molecular Phylogenetics and Evolution [1].

My first impression was one of marvel at the scope of the study I had imposed myself into.  Holy trash snails, Batman!  1,722 individual Galba analyzed from 161 sites in ten countries and territories, over a period of twenty years!  In addition to the 9 populations I myself had contributed (MI, OH, PA, NY, TN, VA, NC, SC-humilis, SC-cubensis), I was especially gratified to discover in the database 5 populations from New Mexico, 6 from Louisiana, 1 from Florida, 1 from Texas, and 2 from way up near Montreal.

This huge sample was first characterized by “shell morphology and reproductive anatomy,” but not by radula, which has been a source of chronic irritation to me for at least ten years now [7], but it’s beginning to look like I’ll have to let that go.  The sample of 1,420 snails not distinguished morphologically was then analyzed using our multiplex PCR technique, about which I also have misgivings, but having dwelled upon those at considerable length last month [22June21], will also set aside at this point.

From Bargues et al. [8]

A subset of the samples passing through multiplex PCR screening were analyzed by a third level of genetic characterization, direct sequencing.  Pili ultimately sequenced 151 individual Galba, mostly the nuclear ITS2 gene and the COI mitochondrial gene, with a few ITS1 and 16S sequences as well, for a total of 300 sequences.  To these 300 sequences she added 517 sequences fished from GenBank “apparently attributable to lymnaeids of the Genus Galba” as follows: 166 COI, 163 ITS2, 118 ITS1, and 70 16S sequences, from 132 New World sites and 45 Old World sites.  The Old World sample was sequenced from 14 countries, the New World sample included 7 countries not sampled in our original coverage.  This stupendous dose of sequence data – which ultimately (setting aside obvious misidentifications) totaled 796 sequences, formed the basis of the analysis we will review this month.

And I feel as though my influence was felt, to some extent, in the priority given to type localities by Pili and her research group.  In Table 1 we listed, in some cases designating for the first time ever, type localities for all eight nomina ultimately playing some role in this research: cousini, cubensis, humilis, meridensis, neotropica, schirazensis, truncatula and viator.  Do look back at the first post in this series [7June21] to refresh your memory on the biology of these eight taxa in particular.  Then for each of these eight nominal species an effort was made to designate type sequences for ITS1, ITS2, 16S and COI, to the extent possible.

But now might be a good opportunity to emphasize that my contribution to the overall research project under review here was much, much less than 1/20 = 0.05 of the effort that ultimately brought the paper to publication.  And as must inevitably be the case in any collaboration of this breadth and complexity, there are elements of the final product of which I am proud, and elements of which I am not-so-proud.

So by all means, feel free to read our introduction, which is the usual boilerplate about how the study that follows will revolutionize worldwide understanding of evolutionary science and cure a plague on the health of the third world.  And by all means, continue through methods sections 2.1 – 2.3, Figure 1 and Table 1.  Then as a personal favor to me, set our paper aside.  Please do not read the remainder of the methods section, or any of the results, or any of the discussion.  Please, I’m begging you.

From Alda et al [1].  Click for larger.

Instead, just look at the four unrooted networks above, inferred by a Baysian technique aptly called “Beast2.”  This figure was composed for a May 2019 draft of our manuscript, but ultimately split into four separate figures for publication [9].  Six colorful blobs have been marked in all four of the networks, as follows:

• The green cluster contains the type population of L. truncatula and 102 other populations collected from Europe and South America.  Good, that’s clear.
• The blue cluster contains the type population of L. humilis from New York, the type populations of the later-described modicella, obrussa, parva, and exigua, and 33 other fossarine populations from the United States and Canada.  These populations are united by shells bearing higher spires and more incised whorls, and by tricuspid first lateral teeth on their radula.  Super!  We're done with all that ancient Baker/Burch mess [10], finally.
• The purple cluster contains the type populations of L. cousini, the more recently-described L. meridensis, and 35 other wide-apertured, sexually-reproducing populations of South America.  Fine, that’s clear as well.
• The yellow cluster contains the type population of L. schirazensis and 84 other fossarine populations from Asia and The Americas.  That meaning of that result is not clear at all, and we will return to it next month.
• The salmon-colored cluster contains the type population of L. cubensis, the more recently-described L. neotropica, and 114 other fossarine populations collected throughout the Americas.  The population of L. cubensis I collected here in the Charleston area is in that salmon blob, as well as a bunch collected from Florida, Louisiana, Texas and New Mexico.  Also included was one 16S sequence from Oklahoma labeled “Fossaria bulimoides” fished from Genbank [11] [20].  These Galba populations bear bicuspid first laterals (as far as is known) and shells with lower spires and more convex whorls.  Well, okay, but…
• The fuchsia cluster contains the type population of L. viator from Argentina and 17 other South American populations.  And good grief.  That fuchsia cluster is contained entirely within the salmon-colored L. cubensis cluster in three of the four networks.

There is no difference of any sort – genetic, morphological, or ecological – between the populations of crappy-little amphibious lymnaeids that d’Orbigny described as Lymnoeus viator in 1835 and the populations of crappy-little amphibious lymnaeids that Pfeiffer described as Limnaea cubensis in 1839.  All have shells bearing more rounded whorls with less incised sutures and bicuspid first lateral teeth on the radula.  The minor ITS1 sequence distinction might simply reflect geography.  Our multiplex PCR results [6] are of no help because they assume the result.  Pili selected her primer mix to amplify bands on a population of nominal cubensis but not on a population of nominal viator.  So our multiplex PCR results cannot subsequently be used to test the hypothesis that cubensis and viator are distinct.

The biological species concept is voided here by asexual reproduction.  The morphological species concept would demand that Pfeiffer’s 1839 cubensis be synonymized under d’Orbigny’s 1835 viator, since the two taxa are indistinguishable.  And from the standpoint of service to humanity, which is a very unfamiliar point for me to stand, some medical and veterinary benefit might be gained by calling all those populations of crappy-little fascioliasis vectors the same thing, rather than two different things, implying that there might be some difference in the best way to control them, which there ain’t.

But darn it.  As I have pointed out on this blog at least a couple times a year for 20 years, the binomial system of nomenclature was not proposed to reflect biological, reproductive, or evolutionary relationships among organisms.  It was proposed for the cataloguing and retrieval of information, a sort-of Dewey Decimal system for critters.  And the first commandment in the Worldview of Information is, Thou Shalt Not Lose Information [12].  And large and important literatures are now indexed under both the names “cubensis” and “viator,” which science now calls us to unite, without losing information under either tab.

North American detail from Fig 2 of [1]

OK, I’m going rogue.  Or roguer than normal, anyway.  In Figure 2 of our MP&E paper (the Galba distribution maps, modified above) we referred to the entire cluster ranging from Carolina to Argentina as “cubensis/viator.”  Henceforth, therefore, the FWGNA Project will refer to crappy-little amphibious lymnaeids with bicuspid first laterals and less-incised shell whorls with the slash-name: “Lymnaea (Galba) cubensis/viator Pfeiffer 1839/d’Orbigny 1835” [12.5].  I am not aware of any law in the international code of zoological nomenclature explicitly prohibiting such an admittedly awkward/vivid Latinate amalgam, but if there is, the FWGNA don’t need no stinking badges [13].  Never have.

Then here on the mudbanks of North American fresh waters, we host two common and widespread species of crappy-little amphibious (“fossarine”) lymnaeids, the tricuspid Lymnaea (Galba) humilis Say 1822 and the bicuspid Lymnaea (Galba) cubensis/viator Pfeiffer 1839/d’Orbigny 1835. Junior synonyms of L. humilis include, but are not limited to: cyclostoma Walker 1908, exigua Lea 1841, galbana Say 1825, modicella Say 1825, obrussa Say 1825, peninsulae Walker 1908, parva Lea 1841, rustica Lea 1841, and tazwelliana Wolf 1869.  Junior synonyms of cubensis/viator include, but are not limited to dalli Baker 1907 and perplexa Baker & Henderson 1929.  Tenuous evidence [11][20] would include bulimoides Lea 1841, cockerelli Pilsbry & Ferris 1906, hendersoni Baker 1909, perpolita Dall 1905, sonomaensis Hemphill 1906, techella Haldeman 1867, and vancouverensis Baker 1939 in the list of cubensis/viator synonyms as well.

Now before we go any further, I feel called to introduce a boxed essay on the concept of “cryptic species.”  Almost all freshwater gastropods are cryptic to almost everybody, I suppose.  Crypsis is in the eye of the beholder.

So the human population of the world is roughly 8 x 10^9.  And the total number of beholders who will ever read the present essay, if I am optimistic, might approach 4 x 10^1.  I am here to re-assure you, the  0.000000005 fraction of the world’s population who constitute my readership, that Lymnaea humilis and Lymnaea cubensis/viator are not entirely cryptic.

The whorls of the shells borne by cubensis/viator are typically more rounded with suture lines less impressed than the whorls of the shells borne by humilis.  Look at Santi Mas-Coma’s figure of the (tricuspid) L. humilis from its New York type locality way up at the top of this essay [5].  Now look at his figure of L. cubensis/viator further down [8].  You, my N = 4 x 10^1 readership, will not usually get populations of those two subsets confused, I feel confident.

Is the shell morphology demonstrated by every snail in every humilis population always distinctive from the shell morphology demonstrated by every snail in every cubensis/viator population?  No, certainly not.  Refer back to Anna Correa’s figure [7] in my essay of [7June21].  But the two Galba species are just as distinctive as Physa acuta and Physa gyrina, or (sometimes) Lymnaea elodes and Lymnaea catascopium [14].  If you, my readership, know what to look for, you will probably make the right identification.

From Baker [15] and Clarke [16]

The reason I am making such a big deal of the shell morphological distinction between humilis and cubensis/viator here is that there is traditionally understood to be a third species of crappy-little amphibious lymnaeid ranging across a narrow, frigid strip of North America, Lymnaea (Galba) truncatula.  The data are shell-morphological only.  We have no genetic evidence one way or the other.

Baker/Burch [10] gave the North American range of L. truncatula as, “portions of Alaska and Yukon Territory.”  Baker himself, however, never laid eyes on an American truncatula.  The shells figured on his plate XXVII were collected from England.

Baker’s American truncatula records seem to have come almost entirely from William Healey Dall [17].  And the challenge is that Dall also recorded populations he identified as Lymnaea (Galba) galbana from Alaska and The Yukon, which we now understand to be a junior synonym of humilis.

Similarly, Arthur Clarke [16] identified truncatula from a small patch of the southern Yukon, as well as from a second patch of southern British Columbia [18] about 1,000 miles south.  But again, Clarke also reported populations of crappy-little amphibious lymnaeids he identified as exigua, ferruginea, modicella, and parva widespread and common throughout the entirety of western Canada, all of which Clarke distinguished minor aspects of shell morphology.  Here’s how Clarke characterized L. truncatula:

“the strongly rounded whorls (shouldered in many specimens), solid shell, deep and partly obscured umbilicus, and general appearance are such distinctive features that identification of this species [truncatula], once seen, can confidently be made.”

To the extent we credit the finely-honed conchological perspicacity of illustrious forefathers such as Dall, Baker, and Clarke, we must admit a third species of crappy-little amphibious lymnaeid to the North American fauna, Lymnaea (Galba) truncatula (Muller 1774).  Viewed from the other side of the Bering Sea, however, Larisa Prozorova [19] listed L. truncatula as a “question mark” in North America.  Her assessment of the situation sounds fair to me.

So here at the end of yet another overly-long essay on yet another justifiably-obscure topic, we find ourselves circling back to the title.  Lymnaea humilis plus L. cubensis/viator certainly sums to two species of Galba inhabiting North America.  Confirmation of Lymnaea truncatula would add a third.

But wait.  Can we back up to that fourth bullet-point way up above?  What’s this business about “The yellow cluster contains the type population of L. schirazensis and 84 other fossarine populations from Asia and The Americas?”  The Americas, pleural?  Might “the infamous schirazensis” complicate our math yet further?  Tune in next time.

Postscript added 9July21 - Our colleague Sam Loker has just published an excellent paper online (open access), figuring shells from 14 North American populations of Galba humilis, including all 8 of the populations I contributed to the Alda et al effort.  See:

• Loker, Dolignow, Pape, Topper, Alda, Pointier, Ebbs, Sanchez, Verocai, DeJong, Brant, and Laidemitt (2021) An outbreak of canine schistosomiasis in Utah: Acquisition of a new snail host (Galba humilis) by Heterobilharzia americana, a pathogenic parasite on the move.  One Health 13: 100280. [html]

Notes:

[1] Alda, Pilar, M. Lounnas, A.Vázquez, R. Ayaqui, M. Calvopiña, M. Celi-Erazo, R.T. Dillon Jr., L. González Ramírez,  E. Loker, J. Muzzio-Aroca, A. Nárvaez, O. Noya, A. Pereira, L. Robles, R. Rodríguez-Hidalgo, N. Uribe, P. David, P. Jarne, J-P. Pointier, & S. Hurtrez-Boussès (2021) Systematics and geographical distribution of Galba species, a group of cryptic and world-wide freshwater snails.  Molecular Phylogenetics and Evolution 157: 107035. [pdf] [html]

[2] The FWGNA Project has adopted the “Hubendick compromise” model for the classification of the Lymnaeidae, recognizing Galba as a subgenus of the worldwide genus Lymnaea [3].  In the series of essays that follows we will often, however, refer to the nomen Galba as though it were a genus, following the usage of the authors whose work we are reviewing.  See:

• The Legacy of Frank Collins Baker [20Nov06]
• The Classification of the Lymnaeidae [28Dec06]

[3] Hubendick, B. (1951)  Recent Lymnaeidae.  Their variation, morphology, taxonomy, nomenclature and distribution.  Kungliga Svenska Vetenskapsakademiens Handlingar Fjarde Serien 3: 1 - 223.

[4] Long-time readers will remember the humilis/cubensis confusion that I resolved by restricting Say’s type locality to New York.  For the rest of you, see:

• Malacological Mysteries I: The type locality of Lymnaea humilis [25June08]

[5] This figure is taken from my unpublished collaboration with Prof. Dr. Santiago Mas-Coma and his colleagues in Valencia:

• Bargues, M.D., P. Artigas, R.T. Dillon, Jr., and S. Mas-Coma (unpubl) Fascioliasis in North America: Multigenic characterization of a major vector and evaluation of the usefulness of rDNA and mtDNA markers for lymnaeids.

[6]  Alda, Pilar, M. Lounnas, A. Vázquez, R. Ayaqui, M. Calvopiña, M. Celi-Erazo, R. T. Dillon, P. Jarne, E. Loker, F. Pareja, J. Muzzio-Aroca, A. Nárvaez, O. Noya, L. Robles, R. Rodríguez-Hidalgo, N. Uribe, P. David, J-P. Pointier, & S. Hurtrez-Boussès (2018). A new multiplex PCR assay to distinguish among three cryptic Galba species, intermediate hosts of Fasciola hepatica.  Veterinary Parasitology 251: 101-105.  [html]  [PDF].  For a review, see:

• The American Galba: Sex, Wrecks, and Multiplex [22June21]

[7] Correa, A.C., J.S. Escobar, O. Noya, L.E. Velasquez, C. Gonzalez-Ramirez, S. Hurtrez-Bousses & J-P. Pointier (2011)  Morphological and molecular characterization of Neotropic Lymnaeidae (Gastropoda: Lymnaeoidea), vectors of fasciolosis.  Infection, Genetics and Evolution 11: 1978-1988.  For a review, see:

• The Lymnaeidae 2012: Fossarine Football [7Aug12]

[8] This is figure 7 from Bargues, M.D., J.B. Malandrini, P. Artigas, C.C. Soria, J.N. Velasquez, S. Camevale, L. Mateo, M. Khoubbane, and S. Mas-Coma (2016)  Human fascioliasis endemic areas in Argentina: multigene characterization of the lymnaeid vectors and climatic-environmental assessment of the transmission pattern.  Parasites and Vectors 9:306.  DOI 10.1186/s13071-016-1589-z

[9] Figures S4 – S7 in the supplementary material ultimately published show painfully-detailed tree-form phylogenies for each of these four genes separately if you are curious about the fine structure.

[10] This is a difficult work to cite.  J. B. Burch's North American Freshwater Snails was published in three different ways.  It was initially commissioned as an identification manual by the US EPA and published by the agency in 1982.  It was also serially published in the journal Walkerana (1980, 1982, 1988) and finally as stand-alone volume in 1989 (Malacological Publications, Hamburg, MI).

[11] Our synonymization of bulimoides Lea 1841 under cubensis/viator is especially tentative [20].  Hubendick [3] considered the nomen valid, to distinguish a rotundly-shelled fossarine ranging from the Mississippi River broadly across the American west.  I have no personal observations one way or the other.  I am quite certain, however, that the single 16S sequence uploaded to GenBank by Remigio, labelled “Fossaria bulimoides” but collected 2,000 miles from the bulimoides type locality in Oregon, is weak evidence, indeed.

• Remigio, E., 2002. Molecular phylogenetic relationships in the aquatic snail genus Lymnaea, the intermediate host of the causative agent of fascioliasis: insights from broader taxon sampling. Parasitol. Res. 88, 687–696.

[12] I developed the idea of duality between the Worldview of Information and the Worldview of Science here:

• When Worlds Collide: Lumpers and splitters [4Sept12]

[12.5] Shortly after this essay was posted, I received a comment from our good friend Bryan England inquiring why I listed cubensis before viator in the slash-construction.  The short answer is that Pfeiffer’s cubensis is the more familiar of the two names here in North America.  But if other workers would prefer to list d’Orbigny’s viator first, they should feel free.  Or by all means, use the single-slash construction that Bryan suggested, rather than the double-slash.  I am not advocating the addition of any additional rules to the code!

[13] Although an optional FWGNA badge has just recently become available.  See:

• Fun With Campeloma! [7May21]

[14] The shell morphology demonstrated by populations of the two big "stagnicoline" lymnaeids we host here in North America, L. elodes and L. catascopium, is very much a function of ecophenotypic plasticity: skinny in the weeds, fat in exposed environments.  Sometimes the two species look different, and sometimes they do not.  This was the subject of an elaborate series on posts from April to July of 2012, and again from June to September of 2015.  Here's the bottom line:

• The Lost Thesis of Samantha Flowers [3Sept15]

[15] Baker, F. C. (1911) The Lymnaeidae of North and Middle America, Recent and Fossil. Special Publication, no. 3. Chicago: Chicago Academy of Natural Sciences.

[16] Clarke, A. (1981) The Freshwater Mollusks of Canada. Ottawa: The National Museums of Canada

[17] Dall, W. (1905) Land and Fresh Water Mollusks of Alaska and Adjoining Regions. Harriman Alaska Expeditions 1899, no. 13. Washington, D.C.: Smithsonian Institution.

[18] Jacquie Lee did not confirm L. truncatula in British Columbia, however, identifying only galbana, modicella, and parva.  See:

• Lee, J.S. (2000)  The distribution and ecology of the freshwater molluscs of Northern British Columbia.  M.Sc. thesis, University of Northern British Columbia, 238 pp.

[19] Prozorova, L. A., 1998. Annotated list of Beringian freshwater mollusks. The Bulletin of the Russian Far Eastern Malacological Society 2: 12–28.  See her Appendix 1.

[20] Note added 13Feb24.  The single 16S sequence in GenBank labeled “Fossaria bulimoides” has proven to be spurious.  And L. bulimoides (Lea 1841) is not a junior synonym of L. cubensis/viator, nor are cockerelli, hendersoni, perpolita, sonomaensis, techella or vancouverensis.  For more, see:

• What is Lymnaea bulimoides? [13Feb24]