Daily Rules, Proposed Rules, and Notices of the Federal Government


Fish and Wildlife Service

50 CFR Part 17

[Docket No. FWS-R3-ES-2012-0065; 4500030113]

RIN 1018-AY16

Endangered and Threatened Wildlife and Plants; Endangered Status for Grotto Sculpin and Designation of Critical Habitat

AGENCY: Fish and Wildlife Service, Interior.
ACTION: Proposed rule.
SUMMARY: We, the U.S. Fish and Wildlife Service, propose to list the grotto sculpin (Cottussp. nov.) as an endangered species under the Endangered Species Act of 1973, and propose to designate critical habitat for the species. In total, all underground aquatic habitat underlying approximately 94 square kilometers (36 square miles) plus 31 kilometers (19.2 miles) of surface stream are being proposed for designation as critical habitat. The proposed critical habitat is located in Perry County, Missouri. If adopted, the effect of these regulations is to conserve grotto sculpin and its habitat under the Endangered Species Act.
DATES: Written Comments:We will accept comments received or postmarked on or before November 26, 2012. Comments submitted electronically using the Federal eRulemaking Portal (seeADDRESSESsection, below) must be received by 11:59 p.m. Eastern Time on the closing date. We must receive requests for public hearings, in writing, at the address shown in theFOR FURTHER INFORMATION CONTACTsection by November 13, 2012.

Public Meeting:To better inform the public of the implications of the proposed listing and proposed critical habitat, and to answer any questions regarding this proposed rule, we plan to hold a public meeting on Tuesday, October 30 from 5-8 p.m. at the Perryville Higher Education Center, 108 South Progress Drive, Perryville, MO 63775.

ADDRESSES: (1)Electronically:Go to the Federal eRulemaking Portal: the Search box, enter Docket No. FWS-R3-ES-2012-0065, which is the docket number for this rulemaking. Then, click the Search button. You may submit a comment by clicking on "Comment Now!." If your comments will fit in the provided comment box, please use this feature of,as it is most compatible with our comment review procedures. If you attach your comments as a separate document, our preferred file format is Microsoft Word. If you attach multiple comments (such as form letters), our preferred format is a spreadsheet in Microsoft Excel.

(2)By hard copy:Submit by U.S. mail or hand-delivery to: Public Comments Processing, Attn: FWS-R3-ES-2012-0065; Division of Policy and Directives Management; U.S. Fish and Wildlife Service; 4401 N. Fairfax Drive, MS 2042-PDM; Arlington, VA 22203.

We request that you send comments only by the methods described above. We will post all comments on generally means that we will post any personal information you provide us (see theInformation Requestedsection below for more information).

The coordinates or plot points or both from which the critical habitat maps are generated are included in the administrative record for this rulemaking and are available at, www.regulations.govat Docket No. FWS-R3-ES-2012-0065, and at the Columbia Missouri Ecological Services Field Office (seeFOR FURTHER INFORMATION CONTACT). Any additional tools or supporting information that we may develop for this rulemaking will also be available at the Fish and Wildlife Service Web site and Field Office set out above, and may also be included in the preamble and/or

FOR FURTHER INFORMATION CONTACT: Amy Salveter, Field Supervisor, U.S. Fish and Wildlife Service, Columbia Missouri Ecological Services Field Office, 101 Park De Ville Drive, Suite A, Columbia, MO 65203; by telephone 573-234-2132; or by facsimile 573-234-2181. Persons who use a telecommunications device for the deaf (TDD) may call the Federal Information Relay Service (FIRS) at 800-877-8339.

This document consists of: (1) A proposed rule to list the grotto sculpin as an endangered species; and (2) a proposed critical habitat designation for the grotto sculpin.

Executive Summary

Why we need to publish a rule.A species may warrant protection through listing under the Endangered Species Act (Act) if it meets the definition of an endangered or threatened species throughout all or a significant portion of its range. This species has been a candidate for listing since 2002, but was precluded from listing by other higher priority actions. The grotto sculpin currently is afforded no protection under the Act, and, because of continued threats, it warrants the protections afforded by listing under the Act. We are proposing to list the grotto sculpin as an endangered species. Listing a species as an endangered species or threatened species and designating critical habitat can only be done by issuing a rule.

The basis for our action.Under the Act, we can determine that a species is an endangered or threatened species based on any of five factors: (A) The present or threatened destruction, modification, or curtailment of its habitat or range; (B) overutilization for commercial, recreational, scientific, or educational purposes; (C) disease or predation; (D) the inadequacy of existing regulatory mechanisms; or (E) other natural or manmade factors affecting its continued existence. We have determined the threats to the species include:

• Habitat loss and degradation of aquatic resources, including such things as illegal waste disposal, chemical leaching, contaminated groundwater, vertical drains, urban development, sedimentation, and industrial sand mining.

• Predation by nonnative predators.

• Inadequate existing regulatory mechanisms that allow significant threats such as water contamination and exploitation of sinkholes.

• Other natural or manmade factors, including loss of genetic diversity, natural environmental variability, and climate conditions such as drought.

This rule proposes to designate critical habitat for the species.If prudent and determinable, we must designate critical habitat for endangered or threatened species. We are required to base the designation on the best available scientific data after taking into consideration economic and other impacts. We can exclude an area from critical habitat if the benefits of exclusion outweigh the benefits of designation, unless the exclusion will result in the extinction of the species. We are proposing to designate critical habitat in Perry County, Missouri, as follows:

• Two units comprised of all underground aquatic habitat underlying approximately 94 km2(36.28 mi2).

• Two units that include approximately 31 kilometers (19.2 miles) of surface stream.

We are preparing an economic analysis.To ensure that we consider the economic impacts, we are preparing an economic analysis of the proposed designation.

We will seek peer review.We are seeking comments from independent specialists to ensure that our listing determination and critical habitat designation are based on scientifically sound data and analyses. We will invite these peer reviewers to comment, during the comment period, on our proposed listing and critical habitat designation. Because we will consider all comments and information received during the comment period, our final determination may differ from this proposal.

Information Requested

We intend that any final action resulting from this proposed rule will be based on the best scientific and commercial data available and be as accurate and as effective as possible. Therefore, we request comments or information from other concerned governmental agencies, Native American tribes, the scientific community, industry, or any other interested parties concerning this proposed rule. We particularly seek comments concerning:

(1) The species' biology, range, and population trends, including:

(a) Habitat requirements for feeding, breeding, and sheltering;

(b) Genetics and taxonomy;

(c) Historical and current range including distribution patterns;

(d) Historical and current population levels, and current and projected trends; and

(e) Past and ongoing conservation measures for the species, its habitat or both.

(2) The factors that are the basis for making a listing determination for a species under section 4(a) of the Act (16 U.S.C. 1531et seq.), which are:

(a) The present or threatened destruction, modification, or curtailment of its habitat or range;

(b) Overutilization for commercial, recreational, scientific, or educational purposes;

(c) Disease or predation;

(d) The inadequacy of existing regulatory mechanisms; or

(e) Other natural or manmade factors affecting its continued existence.

(3) Biological, commercial trade, or other relevant data concerning any threats (or lack thereof) to this species and existing regulations that may be addressing those threats;

(4) Additional information concerning the historical and current status, range, distribution, and population size of this species, including the locations of any additional populations of this species;

(5) The reasons why we should or should not designate habitat as “critical habitat” under section 4 of the Act (16 U.S.C. 1531et seq.), including whether there are threats to the species from human activity, the degree of which can be expected to increase due to the designation, and whether that increase in threats outweighs the benefit of designation such that the designation of critical habitat is not prudent.

(6) Specific information on:

(a) The amount and distribution of grotto sculpin and its habitat,

(b) What may constitute “physical or biological features essential to the conservation of the species,” within the geographical range currently occupied by the species;

(c) Where these features are currently found,

(d) Whether any of these features may require special management considerations or protection;

(e) What areas, that were occupied at the time of listing (or are currently occupied) and that contain features essential to the conservation of the species, should be included in the designation and why,

(f) What areas not occupied at the time of listing are essential for the conservation of the species and why;

(7) Land use designations and current or planned activities in the areas occupied by the species or proposed to be designated as critical habitat, and possible impacts of these activities on this species and proposed critical habitat;

(8) Information on the projected and reasonably likely impacts of climate change on the grotto sculpin and proposed critical habitat;

(9) Any foreseeable economic, national security, or other relevant impacts that may result from designating any area that may be included in the final designation. We are particularly interested in any impacts on small entities, and the benefits of including or excluding areas from the proposed designation that are subject to these impacts;

(10) Whether our approach to designating critical habitat could be improved or modified in any way to provide for greater public participation and understanding, or to assist us in accommodating public concerns and comments;

(11) The likelihood of adverse social reactions to the designation of critical habitat and how the consequences of such reactions, if likely to occur, would relate to the conservation and regulatory benefits of the proposed critical habitat designation.

Please note that submissions merely stating support for or opposition to the action under consideration without providing supporting information, although noted, will not be considered in making a determination, as section 4(b)(1)(A) of the Act directs that determinations as to whether any species is a threatened or endangered species must be made “solely on the basis of the best scientific and commercial data available.”

You may submit your comments and materials concerning this proposed rule by one of the methods listed in theADDRESSESsection. We request that you send comments only by the methods described in theADDRESSESsection.

If you submit information via,your entire submission—including any personal identifying information—will be posted on the Web site. If your submission is made via a hardcopy that includes personal identifying information, you may request at the top of your document that we withhold this information from public review. However, we cannot guarantee that we will be able to do so. We will post all hardcopy submissions on include sufficient information with your comments to allow us to verify any scientific or commercial information you include.

Comments and materials we receive, as well as supporting documentation we used in preparing this proposed rule, will be available for public inspection on,or by appointment, during normal business hours, at the U.S. Fish and Wildlife Service, Columbia Missouri Ecological Services Field Office (seeFOR FURTHER INFORMATION CONTACT).

Public Meeting:We have scheduled a public meeting to be held on Thursday, October 11, 2012 at the Perryville Higher Education Center, 108 South Progress Drive, Perryville, MO 63775. Any interested individuals or potentially affected parties seeking additional information on the public meeting should contact the Columbia Missouri Ecological Services Field Office (SeeFOR FURTHER INFORMATION CONTACT). The U.S. Fish and Wildlife Service is committed to providing access to this event for all participants. Please direct all requests for interpreters, close captioning, or other accommodation to the Columbia Missouri Ecological Services Field Office (SeeFOR FURTHER INFORMATION CONTACT) by 5 p.m. on October 4, 2012.

Previous Federal Actions

We first identified the grotto sculpin as a candidate species in a notice of review published in theFederal Registeron June 13, 2002 (67 FR 40657). Candidate species are assigned listing priority numbers (LPNs) based on the immediacy and magnitude of threats, as well as taxonomic status. The lower the LPN, the higher priority that species is for us to determine appropriate action using our available resources. The grotto sculpin was assigned an LPN of 2 due to imminent threats of a high magnitude. On May 11, 2004, we received a petition dated May 4, 2004, from The Center for Biological Diversity to list 225 candidate species, including the grotto sculpin. From 2004 through 2011, notices of review published in theFederal Register(69 FR 24876, 70 FR 24870, 71 FR 53756, 72 FR 69034, 73 FR 75176, 74 FR 57804, 75 FR 69222, 76 FR 66370) continued to maintain an LPN of 2 for the species.

Status Assessment for Grotto Sculpin Background Species Description

The grotto sculpin (Cottussp. nov.) is a cave-dwelling fish that exhibits characteristics typical of troglomorphic (adapted to living in constant darkness) organisms, including greatly reduced or absent eyes and skin pigmentation (Burret al.2001, p. 286). The grotto sculpin is moderately-sized relative to other species in the genus; the largest specimen examined by Adamset al.(unpub. data) was 104 millimeters (mm) (4.1 inches (in)) standard length (SL).


The grotto sculpin belongs to the family Cottidae (Pflieger 1997, p. 253) and until recently was considered to be a member of the banded sculpin (Cottus carolinae) complex. The banded sculpin occurs in streams and rivers in adjacent watersheds; however no otherCottusoverlaps the geographic range of the grotto sculpin. Burret al.(2001, p. 293) demonstrated that hypogean (underground) grotto sculpin found in Perry County, Missouri, are morphologically distinct from the epigean (above ground) forms of banded sculpin found outside the Cinque Hommes Creek drainage in that they exhibit obvious troglomorphic characteristics and other unique anatomical variations. Although the occurrence of banded sculpin in subterranean waters is well known, none of these sculpin shows evidence of cave adaption exhibited by grotto sculpin, and none is known to be a permanent cave resident. Grotto sculpin are distinguished from all otherCottusspecies, except banded sculpin, by the complete lateral line terminating near the base of the caudal fin and lack of connection between dorsal fins (Adamset al.unpub. data). The grotto sculpin is distinct from the banded sculpin based on a reduction in eye size and an increase in cephalic lateralis pore size (Adamset al.unpub. data). Morphology of brain structures in hypogean individuals also differs significantly from that of epigean banded sculpin, including reduced optic and olfactory lobes and enlarged inferior lobe of the hypothalamus, eminentia granularis, and crista cerebellaris (Adams 2005, pp. 17-18).

Population genetics ofCottussculpin in southeast Missouri also have been analyzed. Adamset al.(unpub. data) conducted a population genetics study of sculpin from the Bois Brule drainage in Perry County, the Greasy Creek in Madison County, and the Current River in Ripley County. Unique evolutionary lineages for each of the three areas, based on distinct nuclear haplotypes, were identified and supported. A single nuclear haplotype was identified among sampled individuals throughout the Bois Brule drainage (Mystery Cave, Running Bull Cave, Rimstone River Cave, Crevice Cave, Moore Cave, and Cinque Hommes Creek), a second from Greasy Creek, and a third from the Current River. Adamset al.(unpub. data) is in the process of formally describing the grotto sculpin as a taxonomically distinct species based on the combination of morphologic and genetic uniqueness. Morphological data alone are not definitive in supporting a unique taxonomic unit; however, morphological data augmented by the results of genetic analyses by Adamset al.(unpub. data) support the divergence of grotto sculpin from otherCottusspecies.

Life History and Habitat

Grotto sculpin occupy cave streams, resurgences (also known as “spring branches”) (Vandike 1985, p. 10), springs, and two surface streams (Adams 2012, pers. comm.;Burret al.2001, p. 284). Resurgences refer to the point of emergence of a cave stream from the cave system and are an interface between strictly subterranean habitats (caves) and streams that flow only on the surface. Age-class distribution of grotto sculpin between cave and surface habitats shifts throughout the year, but in general, adults make up a higher percentage of overall grotto sculpin abundance in caves, whereas juveniles comprise a higher percentage of overall abundance on the surface (Gerken 2007, p. 14). Adults increase in abundance at resurgence sites in October, peak in December, and disappear from resurgence sites in January (Adamset al.2008, p. 5). Such seasonal changes in adult abundance might be indicative of a subterranean migration for spawning (Adams 2005, p. 50).

The appearance of grotto sculpin young-of-year in spring and early summer suggests late winter and early spring spawning (Day 2008, p. 18). The distance grotto sculpin travel upstream in caves is unknown, but a nest has been observed 0.6 meters (m) (2 feet (ft)) inside the cave portal at Thunderhole Resurgence, indicating they might stay close to surface habitats (Adamset al.2008, p. 8). Five nests, with approximately 200 eggs each, were discovered within a 100-m (328-ft) area in Mystery Cave in December 1998, suggesting synchronous spawning within the cave (Adams 2005, p. 10). Nests were adhered to the underside of rocks in flowing water with a temperature of 14 °C (57 °F). Reproduction could occur as late as February or March in Cinque Hommes Creek, based on the observation of yolk-sac larvae and a single nest (Adamset al.unpub. data). Spawning could be tied to water temperature, with temperatures reaching optimum levels in caves as early as 2 to 3 months before surface habitats, explaining why spawning was not observed concurrently in those habitats (Adams 2005, pp. 10-11). Males remain present at nests and guard rocks to which nests are attached (Adamset al.unpub. data).

Young-of-year abundance increases between March and May at resurgence sites, and between April and May in caves (Adamset al.2008, p. 5). That increase, coupled with decreased recaptures, likely is a result of young-of-year recruitment into the population. Adamset al.(2008, p. 7) classified grotto sculpin 30 mm (1.2 in) or less in length to be juveniles. At this size they can be tagged but are still susceptible to predation by adult sculpin as well as invasive fish. Grotto sculpin are cannibalistic, with the young providing a potential food source for adults in an otherwise forage-limited environment (Adamset al.2008, p. 7). Seasonal decreases in abundance of young-of-year and juveniles likely are the result of spring and summer predation and cannibalism in addition to other causes of mortality. Epigean fishes, such as green sunfish (Lepomis cyanellus), bluegill (L. macrochirus), and channel catfish (Ictalurus punctatus), can access caves through sinkholes and arepotential predators on eggs and juveniles (Burret al.2001, p. 284).

Resurgences are used by juvenile grotto sculpin as nursery areas, where the juveniles maximize growth before migrating upstream into caves to reproduce or downstream to surface streams (Day 2008, p. 18). As juveniles grow, the potential for cannibalism decreases and mortality rates stabilize, resulting in increased recapture rates in caves. Both growth rate and metabolism are lower in caves versus resurgence sites (Adams 2005, p. 61; Adamset al.2008, p. 8). However, fish in both habitats reach comparable lengths, alluding to greater longevity of fish in caves (Adamset al.2008, p. 8).

Grotto sculpin tend to occur singly or in small aggregations of 2 to 3 individuals and can be found in the open water or hidden under rocks (Burret al.2001, p. 284). They occupy pools and riffles with moderate flows and variable depths (4 to 33 centimeters (cm) (1.6 to 13 in)) (Burret al.2001, p. 284). Although grotto sculpin have been documented to occur over a variety of substrates (for example, silt, gravel, cobble, rock rubble, and bedrock), the presence of cobble or pebble is necessary for spawning (Burret al.2001, p. 284; Adamset al.unpub. data). Gerken (2007, p. 16) examined habitat use by grotto sculpin in Mystery and Running Bull caves, Cinque Hommes Creek, and Thunderhole Resurgence. Grotto sculpin tend to be associated with a high availability of invertebrate prey, deeper cave pools, substrate containing cobble, and some level of sustained water flow (Gerken 2007, pp. 16-17). Use of surface habitat by grotto sculpin is most influenced by an abundance of amphipods and isopods. When surface streams with fewer prey items were used, available habitat was more than 23 percent clay. Grotto sculpin in caves occupied deeper pools where cobble comprised at least 10 percent of available habitat, and where amphipods and isopods were in greater abundance. Lower abundances of grotto sculpin were found in shallow cave pools where the substrate consisted of silt deposits deeper than 1.9 cm (0.8 in) (Gerken 2007, p. 16). Silt covered more overall area of available cave habitat, and silt also was deeper in caves compared to surface sites (Gerken and Adams 2007, p. 76).

Within and among caves and streams, sculpin typically move 0 to 50 m (0 to 164 ft) (Adamset al.2008, p. 6). Over multiple sampling trips, substantial migrations greater than 200 m (656 ft) have been observed (range 0 to 830 m (0 to 2,723 ft)). The largest single movement of sculpin observed between two subsequent sampling trips (October to December 2007) was 610 m (2,001 ft) in Mystery Cave (Adamset al.2008, p. 8). Such movements are seasonal and likely related to spawning and avoidance behavior of juveniles to escape predation by adult sculpin (Adamset al.2008, p. 7). In May 2008, an individual that was tagged previously in Running Bull Cave was recaptured in Thunderhole Resurgence, evidencing the physical and biological connection of these two systems (Adamset al.2008, p. 8).

Species Distribution and Status

The grotto sculpin was first documented in 1991 (Adams 2005, p. 11). Burret al.(2001, pp. 280, 284) explored caves in five states that had extensive areas of karst to delineate the geographic range of the grotto sculpin, but found them to exist only in Missouri. Nine karst areas in Perry County, Missouri, were searched because sculpin (Cottussp.) were previously known to be present in those areas, and the karst geology in those nine areas could provide suitable habitat for the grotto sculpin. Based on that study, the grotto sculpin is currently restricted to two karst areas (limestone regions characterized by sink holes, abrupt ridges, caves, and underground streams) in Perry County, Missouri: Central Perryville and Mystery-Rimstone (Burret al.2001, p. 283). Cave systems such as these that form beneath a sinkhole plain provide substantial organic input and an abundance of invertebrates. Such systems might be the only habitats that provide sufficient food and sustained water flow to support grotto sculpin populations (Burret al.2001, p. 291; Day 2008, pp. 16-17). Peck and Lewis (1978, pp. 43-53) documented an abundance of potential prey items in the karst region of southeast Missouri, including isopods, amphipods, flatworms, and snails.

The grotto sculpinis restricted to Blue Spring Branch (from the Moore Cave System resurgence to the confluence with Bois Brule Creek) and the Cinque Hommes Creek drainage, including underlying caves and Cinque Hommes Creek, its tributaries, resurgences, and springs. Within the Cinque Hommes Creek drainage, populations have been documented in five cave systems: Moore Cave, Crevice Cave, Mystery Cave, Rimstone River Cave, and Running Bull Cave (Adamset al.unpub. data; Adams 2012, pers. comm.). Within these cave systems, grotto sculpin occur in cave streams and associated resurgences and springs. Cinque Hommes Creek and Blue Spring Branch are the only surface streams where grotto sculpin have been found. Cinque Hommes Creek is the primary resurgence stream for caves in the Mystery-Rimstone Karst and Crevice Cave in the Central Perryville Karst, whereas Blue Spring Branch is the resurgence stream for the Moore Cave System (Burret al.2001, p. 284). To date, over 153 additional caves in Arkansas, Illinois, Indiana, Missouri, and Tennessee have been searched for grotto sculpin and epigean or hypogean forms of banded sculpin. Of these, banded sculpin was documented in 25 caves, but only fish in the Central Perryville and Mystery-Rimstone karst areas exhibited the cave adaptations characteristic of grotto sculpin (Burret al.2001, p. 284). The full extent of the species' range is unknown because not all reaches in occupied cave systems can be accessed and not all potential, suitable caves, springs, and surface streams have been surveyed (for example, Keyhole Spring; Moss and Pobst 2010, p. 152). We consider the geographic range of the grotto sculpin to be the extent of the Central Perryville and Mystery-Rimstone karst areas, which encompass approximately 222 km2(89 mi2) (Service 2012 calculations based on Burret al.2001, p. 282 and Vandike 1985, p. 1).

There are no total population estimates for the grotto sculpin. Mystery (MC) and Running Bull (RBC) caves and their associated resurgence streams, Mystery Resurgence (MR) and Thunderhole Resurgence (TR), respectively, apparently have the largest populations of grotto sculpin (Adamset al.2008, p. 4). A study conducted from August 2005 to October 2008 yielded a total of 6,265 captures (4,218 individuals) at those four sites (Day 2008, p. 12). The 2,684 (43 percent) captures in caves represented 1,642 individuals, whereas 3,581 (57 percent) captures in resurgences represented 2,576 individuals (Day 2008, pp. 13, 15). Of the captured fish, 2,986 (MC-894, RBC-154, MR-376, TR-1562) were tagged for a mark-recapture study. Mean recapture was higher in caves (46 percent) than resurgences (18 percent) (Day 2008, p. 13). Grotto sculpin densities were significantly lower in caves (0.037/m2(0.398/ft2)) compared to resurgence streams (0.225/m2(2.42/ft2)) (Day 2008, p. 13). Density at Thunderhole Resurgence was significantly higher (0.610/m2(6.57/ft2)) than any other site surveyed (MC 0.036/m2(0.388/ft2), RBC 0.113/m2(1.22/ft2), MR 0.032/m2(0.344/ft2)).

Capture success, recapture rates, and population density differ seasonally. The greatest number of grotto sculpin has been captured in summer, followedby spring, fall, and winter (Adamset al.2008, p. 5; Day 2008, p. 12). Overall recapture rates were highest in fall and winter (32 percent each) and lower in spring (25 percent) and summer (15 percent). Overall recapture rates also were significantly lower at resurgence sites than caves, regardless of season. Recapture rates at caves were highest in winter (52 percent) and lowest in fall (44 percent). Recapture rates at resurgence sites were highest in spring (15 percent) and lowest in winter (7 percent). Similar patterns of seasonal changes in density were observed in caves and resurgences. In both habitats, densities were highest in summer, nearly equal in fall and spring, and lowest in winter (Adamset al.2008, p. 5).

Two mass mortalities of grotto sculpin have been documented in Perry County. The first occurred in Running Bull Cave in 2001, when the population was completely lost (Burret al.2001, p. 294; Adams 2005, p. 40). The second occurred in Mystery Cave in August 2005, and affected the uppermost 690 m (2,264 ft) of cave stream (Adamset al.2008, p. 6). Both events were thought to have been caused by point-source pollution (Burret al.2001, p. 294; Adamset al.2008, p. 6). Both caves were recolonized following the die-offs, and grotto sculpin were captured 2 years after the mortality event in Running Bull Cave (Adamset al.2003, p. 7). Surveys were conducted as part of a research study immediately following the die-off in Mystery Cave (Adamset al.2008, p. 6). From August 2005 through March 2006, no grotto sculpin were captured in the upstream sections of Mystery Cave. The first capture of a grotto sculpin after the die-off occurred in May 2006. The first recaptures of three individuals from three different stream sections (540, 560, and 570 m (1772, 1837, and 1870 ft)) occurred in July 2006. Stream sections that supported the earliest recolonization of grotto sculpin in the upper sections (0 to 690 m (0 to 2264 ft)) of Mystery Cave were the most downstream portion of the stream in which the die-off occurred (sections farthest away from the source of contamination). The grotto sculpin population in Mystery Cave increased over the next 3 years to more than 60 individuals in 2007 (Adamset al.2008, p. 8).

Summary of Factors Affecting the Species

Section 4 of the Act (16 U.S.C. 1533), and its implementing regulations at 50 CFR part 424, set forth the procedures for adding species to the Federal Lists of Endangered and Threatened Wildlife and Plants. Under section 4(a)(1) of the Act, we may list a species based on any of the following five factors: (A) The present or threatened destruction, modification, or curtailment of its habitat or range; (B) overutilization for commercial, recreational, scientific, or educational purposes; (C) disease or predation; (D) the inadequacy of existing regulatory mechanisms; and (E) other natural or manmade factors affecting its continued existence. Listing actions may be warranted based on any of the above threat factors, singly or in combination. Each of these factors is discussed below.

Factor A. The Present or Threatened Destruction, Modification, or Curtailment of Its Habitat or Range

The grotto sculpin is a cave-adapted species that is endemic to karst habitats that provide consistent water flow, high organic input, and connection to surface streams, which allow for seasonal migrations to complete its life cycle. Nearly all of the land within the known range of the grotto sculpin is privately owned. Two exceptions are Ball Mill Resurgence Natural Area (19.5 ac (7.9 ha)) and Keyhole Spring and Resurgence near Blue Spring Branch; both properties are owned by the L-A-D Foundation (a private foundation dedicated to sustainable forest management and protection of natural and cultural areas in Missouri ( and managed by the Missouri Department of Conservation (MDC)). The municipality of Perryville is in the Central Perryville Karst Area and is within the recharge area of Crevice Cave. Thirty-six percent (15.6 km2(6.02 mi2)) of Perryville's total area of 43 km2(16.6 mi2) lies within the karst area, whereas 24 percent (10.4 km2(4.02 mi2)) lies within the southern portion of the recharge area of Crevice Cave (recharge area defined by Moss and Pobst 2012, pp. 151-152).

The karst in Perry County is characterized by thousands of sinkholes (Vandike 1985, p. 1) and over 700 caves (Foxet al.2009, p. 5). Water quality in karst areas is highly vulnerable and can severely decline with rapid transmission of contaminants from the surface to the aquifer (Panno and Kelly 2004, p. 230). Moss and Pobst delineated recharge areas for known and potential grotto sculpin caves (2010, pp. 146-160) and evaluated the vulnerability of groundwater in the recharge areas to contamination (2010, pp. 161-190). Because the grotto sculpin is dependent not only on caves, but uses surface habitat in addition to caves, Moss and Pobst (2010, p. 161) evaluated hazards within and adjacent to recharge areas to best characterize impairment of cave and surface streams. They found all the recharge areas to be highly vulnerable and contain hazards from historical sinkhole dumps, agricultural practices without universal application of best management practices, ineffective private septic systems, and roads with contaminated runoff (Burret al.2001, p. 294; Moss and Pobst 2010, p. 183). They noted additional hazards in the recharge area for Crevice Cave not found elsewhere, such as hazardous waste generators, wastewater outflows, storm water outflows, and underground storage tanks for hazard waste, that compound potential threats to groundwater and drinking water (Moss and Pobst 2010, p. 184). Impacts to groundwater are not proportional to the area impacted in such a highly vulnerable landscape—a localized pollution event can impact all aquatic habitats downstream.

There are approximately 2 sinkholes per km2(6 per mi2) in Perry County and 7 sinkholes per km2(17 per mi2) in the Central Perryville and Mystery-Rimstone karst areas (Missouri Department of Natural Resources 2010, unpaginated). Recharge areas around grotto sculpin caves contain up to four times the number of sinkholes compared to other parts of the county or other karst areas. Cave recharge areas in the Central Perryville Karst contain an average of 8 sinkholes per km2(22 per mi2), whereas those in the Mystery-Rimstone Karst contain an average of 4 per km2(11 per mi2) (Missouri Department of Natural Resources 2010, unpaginated). Water flow in Perry County karst systems occurs by way of surface features, such as sinkholes and losing streams, as well as connectivity to the underlying aquifer (Aley 1976, p. 11; Foxet al.2009, p. 5). Without adequate protection, sinkholes can funnel storm-runoff directly into cave systems in a short period of time (Aley 1976, p. 11; White 2002, p. 88; Foxet al.2010, p. 8838).

Illegal Waste Disposal and Chemical Leaching—At least half of the sinkholes in Perry County have been or are currently used as dump sites for anthropogenic waste (Burret al.2001, p. 294). Although it is illegal to dump waste in open sites in Missouri, the practice continues today—sinkholes continue to be used as dump sites for household wastes, tires, and occasionally dead livestock (;Pobst 2012, pers. comm). Moss and Pobst (2010, p. 169) observed that most historical farms in the sinkhole plain had at least one sinkhole that contained household andfarm waste. Waste material found in sinkholes includes, but is not limited to, household chemicals, sewage, and pesticide and herbicide containers (Burret al.2001, p. 294). Foxet al.(2010, p. 8838) found that Perry County cave streams were contaminated by a mixture of organic pollutants that included both current-use and legacy-use pesticides and their degradation products. They found high concentrations of heptachlor epoxide and trans-chlordane, which are degradation products of the legacy-use pesticides heptachlor and chlordane (Foxet al.2010, p. 8839). Heptachlor and chlordane were banned in 1988, but can persist in the environment through storage in sediments above or below ground or leaking containers in sinkholes (ATSDR 1994a, unpaginated; ATSDR 2007a, unpaginated). In water, heptachlor readily undergoes hydrolysis to a compound, which is then readily processed by microorganisms into heptachlor epoxide (ATSDR 2007b, p. 98). Heptachlor and chlordane are highly persistent in soils, are almost insoluble in water, and will enter surface waters primarily though drift and surface run-off (ATSDR 1994a, unpaginated; ATSDR 2007a, unpaginated). Although not specifically tested on the grotto sculpin, both heptachlor and chlordane are highly toxic to most fish species tested, including warm-water species such as bluegill (Lepomis macrochirus) and fathead minnow (Pimephales promelas) (Johnson and Finley 1980, pp. 19, 43-44). Heptachlor caused degenerative liver lesions, enlargement of the red blood cells, inhibited growth, and mortality in bluegill (Andrewset al.1966, pp. 301-305). Heptachlor, heptachlor epoxide, and chlordane have been shown to bioaccumulate in aquatic organisms such as fish, mollusks, insects, plankton, and algae (ATSDR 1994b, p. 172; ATSDR 2007b, p. 89).

Chemical leaching in sinkholes likely is a major contributor to the occurrence of legacy-use pesticides, such as dieldrin, in aquatic habitats (Foxet al.2010, p. 8840). Dieldrin, a domestic pesticide used in the past to control corn pests and cancelled by the U.S. Department of Agriculture (USDA) in 1970 (ATSDR 2002, unpaginated), was found at levels that exceeded ambient water quality criterion by 17 times in Mertz Cave and Thunderhole Resurgence (Mystery-Rimstone Karst Area) (Foxet al.,p. 8839). Dieldrin is a known endocrine disruptor that bioaccumulates in animal fats, especially those animals that eat other animals and, therefore, is a concern for the grotto sculpin because it is the top predator in its cave habitat (ATSDR 2002, unpaginated; Foxet al.2010, p. 8839). The grotto sculpin depends on several species of cave amphipods, includingGammarussp. (Gerken 2007, pp. 16-17; Foxet al.2010, p. 8839). Dieldrin has been detected in the amphipodG. troglophilusthrough tissue bioassays (Tayloret al.2000, p. 10). Tarzwell and Croswell (1957, pp. 253-255) found that dieldrin was toxic to fathead minnow, bluegill, and green sunfish (Lepomis cyanellus). Whereas the species exhibited differences in susceptibility, individuals of all species tested ultimately experienced loss of equilibrium followed by death (Tarzwell and Croswell 1957, p. 255).

Sinkholes have also been used as disposal sites for dead livestock (Foxet al.2009, p. 6; Moss and Pobst 2010, p. 170). Animal carcasses dumped into sinkholes and cave entrances are potentially diseased and could carry pathogens that could be unintentionally introduced into the groundwater system. Decomposing animals in source water for cave streams also can lower the dissolved oxygen and negatively impact aquatic organisms. One of two documented mass mortalities of the grotto sculpin was likely caused by a dead cow in the surface stream above Mystery Cave (Adams 2012, pers. comm.).

Contaminated Water—In cave streams sampled by Foxet al.(2010, p. 8838), time-weighted average (TWA) water concentrations of 20 chemicals were at levels above method detection limits (MDLs); 16 of the 20 chemicals originated from agricultural pest management activities. Acetochlor, diethatyl-ethyl, atrazine, and desethylatrazine (DEA) were detected at all sites during both May and June sampling periods. Pyrene, metolachlor, DEET, and pentachloroanisole were detected at all sites during sampling periods (Foxet al.2010, p. 8838). There is a long list of potential impacts of these chemicals on fish, including reductions in olfactory sensitivity, immune function, and sex hormone concentrations; endocrine disruption; and increased predation and mortality due to adverse effects to behavior (Alvarez and Fuiman 2005, pp. 229, 239; Rohr and McCoy 2010, p. 30). The ubiquitous presence of current-use pesticides, such as atrazine, was not surprising based on the extensive agricultural land use in Perry County. Atrazine has been the most frequently detected herbicide in ground and surface waters in Perry County (Foxet al.2010, p. 8838) and in a similar karst and agricultural landscape in Boone County, Missouri (Lerch 2011, p. 107); levels of corn production were similar in the two counties. Even at concentrations below U.S. Environmental Protection Agency (EPA) criteria for protection of aquatic life, atrazine has been shown to reduce egg production and cause gonadal abnormalities in fathead minnows (Tillettet al.2010, pp. 8-9). Sex steroid biosynthesis pathways and gonad development in male goldfish (Carassius auratus)were impacted by atrazine in concentrations as low as 1 nanogram per liter (ng/L) (Spanoet al.2004, pp. 367-377). Concentrations of atrazine in Perry County ranged from 20 to 130 ng/L (Foxet al.2010, p. 8838). Liet al.(2009, pp. 90-92) showed that environmentally relevant concentrations of acetochlor can decrease circulating thyroid hormone levels, decrease expression of thyroid hormone-related genes, affect normal larval development, and affect normal brain development. Pyrene is known to cause anemia, neuronal cell death, and peripheral vascular defects in larval fish (Incardonaet al.2003, p. 191). Wanet al.(2006, pp. 57-58) considered metolachlor to be slightly to moderately toxic to freshwater amphibians, crustaceans, and salmonid fishes. Wolf and Moore (2010, pp. 457, 464-465) demonstrated that sublethal concentrations of metolachlor adversely affected the chemosensory behavior of crayfish and likely impacted its ability to locate prey. These researchers also noted that this herbicide also caused physiological impairment that likely impacted locomotory behavior and predator avoidance responses. Due to the importance of chemosensory organs to the grotto sculpin, the presence of metolachlor in occupied streams may impact this fish's ability to locate prey.

Additional potential adverse effects to grotto sculpin from contaminants include increased susceptibility to fish disease (Arkooshet al.1998, p. 188), increased immunosuppression (Arkooshet al.1998, p. 188), disruption of the nervous system by inhibition of cholinesterase (Hill 1995, p. 244), and an increase in acute or chronic stress resulting in reduced reproductive success, alterations in blood and tissue chemistry, diuresis, osmoregulatory dysfunction, and reduction in growth (Wedemeyeret al.1990, pp. 452-453). As a result, potential water contamination from various sources of point and non-point source pollution poses a significant, ongoing threat to the grotto sculpin.

Vertical Drains—Potential contaminant problems with sinkholes are further exacerbated by the presence and continued installation of verticaldrains across the agricultural landscape in Ste. Genevieve and Perry Counties (Perry County Soil and Water Conservation District (PCSWCD) 2012, unpaginated). Vertical drains are also known as “stabilized sinkholes” and are defined by the U.S. Department of Agriculture's Natural Resources Conservation Service (NRCS) as “a well, pipe, pit, or bore in porous, underground strata into which drainage water can be discharged without contaminating groundwater resources” (NRCS 2006, p. 1). This conservation practice is meant to reduce erosion by facilitating drainage of surface or subsurface water. Vertical drains often result in more land available to the farmer. As of 2012, the recharge areas for known and potential grotto sculpin habitat in the Central Perryville and Mystery-Rimstone karst areas contained an average of 2.5 vertical drains per km2(7 per mi2), with the highest concentrations in the recharge areas for Keyhole Spring, Ball Mill Spring, and Mystery Cave (PCSWCD 2012, unpaginated). New vertical drains continue to be installed on the landscape at a rate consistent with the installation rate that occurred in the 1990s, with approximately 40 new vertical drains installed at 15 properties in Perry County in 2011 (PCSWCD 2012, unpaginated).

The NRCS (2006, p. 2) noted that “significant additions to subsurface water sources may raise local water tables or cause undesirable surface discharges down-gradient from the vertical drain.” The impact of vertical drains on groundwater has been studied on a limited basis and studies have directly linked groundwater and drinking water contamination with vertical drains (EPA 1999, unpaginated). According to the conditions set by the NRCS, this practice can only be applied when it will not contaminate groundwater or affect instream habitat by reducing surface water flows (NRCS 2010b, p. 1). The NRCS provides a cost-share of up to 75 percent for installation of vertical drains to stop erosion (NRCS 2010b; 2011; 2012) and has conservation practice and construction standards that include secure placement of the standpipe, appropriate fill material around the drainage pipe, and a filter system around the drain (NRCS 2006a, pp. 1-2; 2006b, pp. 1-3). Without implementation of the suite of standards, vertical drains might allow contaminated water to flow directly into caves without naturally occurring filtration (Pobst and Taylor 2007, p. 69). Vertical drains act as conduits for all surface water, contaminants, and sediment directly from the surface through the bedrock into underground caves, streams, and karst voids (Pobst and Taylor 2007, p. 69). Although USDA requires landowners to install a minimum of 7.62 m (25 ft) of grassed buffer around vertical drains to minimize erosion and the migration of nutrients and contaminants into the groundwater system, this guideline is not strictly followed (Moss and Pobst 2010, p. 170). Because vertical drains are potential targets for illegal dumpling of liquid hazardous wastes (Foxet al.2010, p. 8839) and there is an absence of adequate buffers around some vertical drains, the migration of sediment and contaminants is easily facilitated (Moss and Pobst 2010, p. 171). Such a scenario is supported by Foxet al.'s (2010, pp. 8835-8840) contaminant study in the karst region of Perry County. The long list of harmful chemicals detected in the Foxet al.(2010, pp. 8835-8840) study is likely due to the migration of these contaminants directly from surface fields into the underground karst system through vertical drains and sinkholes.

Urbanization and Development—In addition to contamination from point sources of pollution and improper trash disposal, water quality of sculpin habitats is negatively impacted by urban growth of Perryville, located in the recharge area for Crevice Cave (Moss and Pobst 2010, p. 164). Crevice Cave had the lowest amount of cropland and grassland within its recharge and the most chemical detections. In contrast, Mystery Cave had the most cropland and grassland and fewest chemical detections (Foxet al.2010, p. 8840). The only hazardous waste facility in the Central Perryville and Mystery-Rimstone karst areas is located in Perryville. The facility is permitted by the Missouri Department of Natural Resources as a large-volume hazardous waste generator. Additional hazards in Perryville include four other hazardous waste generators; nine underground storage tanks that could leak petroleum products; two National Pollutant Discharge Elimination System (NPDES) permits for wastewater outfalls; and seven NPDES permits for storm water discharge, leaking sewer lines, or lines that remain plumbed into the caves below (Missouri Department of Natural Resources (MDNR) 2010, unpaginated).

Most of the runoff water in areas that recharge aquatic habitats for the grotto sculpin moves quickly into the groundwater system with ineffective natural filtration, and the same is true for waste waters from septic systems (Aley 2012, pers. comm.). Contamination of groundwater by septic systems in karst areas has been documented on multiple occasions (Simon and Buikema 1997, pp. 387, 395; Pannoet al.2006, p. 60) because septic tank systems are poorly suited to karst landscapes (Aley 1976, p. 12). Panno and Kelly (2004, p. 229) listed septic systems as potential contributors of excess nitrogen to streams in the karst region of southern Illinois. Septic systems in the sinkhole plain can be direct conduits for introduction of septic effluent directly into the shallow karst aquifer (Pannoet al.2001, p. 114). In a karst area in southwest Missouri, poorly designed sewage treatment lagoons were allowing effluent from a small, rural school to seep into the only known location for the federally listed Tumbling Creek cavesnail (Antrobia culveri) (Aley 2003, unpaginated).

Most of the rural residents in the Central Perryville and Mystery-Rimstone karst areas employ on-site septic systems (for example, in the Mystery Cave area) (Aley 1976, p. 12). Failure of septic systems occurs in karst areas of southeast Missouri, such as those in Perry County, but detections are problematic because most failures are not obvious from the surface, but instead occur underground into the groundwater system (Aley 2012, pers. comm.). One instance of a septic system failure was observed by Aley (1976, p. 12) near Mystery Cave. Sewage was being discharged to a septic field within 100 ft (30.5 m) of the cave entrance and was contaminating the waters of the Mystery Cave system. Water samples collected by the Missouri Department of Conservation within the range of the grotto sculpin indicated the presence ofEscherichia coliat high levels, which might correspond to high inputs of phosphorus from septic systems (Pobst 2010, pers. comm.). Tayloret al.(2000, pp. 13-16) found that fecal contamination of karst groundwater is a serious problem in southeast Missouri. Among sampling locations in southeast Missouri, water samples were taken from streams and springs in Perry County that included sites within the range of the grotto sculpin (Mertz Cave, Running Bull Cave, Thunderhole Resurgence, and Cinque Hommes Creek) (Tayloret al.2000, pp. 48-49). High fecal bacterial loads were found in groundwater of grotto sculpin habitats and can be a combination of both human and animal wastes (Tayloret al.2000, p. 14).

No animal feeding operations (AFOs) or concentrated animal feeding operations (CAFOs) are present in the recharge areas of grotto sculpin habitat (MDNR 2010), but there are smaller livestock feeding areas that are in sinkholes or near sinkhole drainagepoints (Aley 1976, p. 12; Moss and Pobst 2010, p. 166). Large amounts of manure can be flushed through sinkholes and carry associated bacteria and pathogens into cave streams. Waste from mammalian sources, including humans and livestock, can increase nutrient loads and lower dissolved oxygen in the groundwater (Simon and Buikema 1997, p. 395; Pannoet al.2006, p. 60). Hypoxia resulting from eutrophication due to increases in nutrient load (especially phosphorus) can lead to mortality and sublethal effects by reducing the availability of oxygen needed by fish for locomotion, growth, and reproduction (Kramer 1987, p. 82; Gould 1989-1990, p. 467), Barton and Taylor (1996, p. 361) reported that low dissolved oxygen levels can cause changes in cardiac function, increased respiratory and metabolic activity, alterations in blood chemistry, mobilization of anaerobic energy pathways, upset in acid-base balance, reduced growth, and decreased swimming capacity of fish.

Sedimentation—Concerns with sedimentation (actual deposition of sediment, not the transport) and wash load (portion of the sediment in transport that is generally finer than the sediment) (as defined by Biedenharnet al.2006, pp. 2-6) relative to impacts to grotto sculpin habitat are primarily the transport of contaminants and the deposition of excessive amounts of sediment in cave streams. Soils in the Central Perryville and Mystery-Rimstone karst areas are dominated by highly erosive loess. Sediment transported into the karst groundwater can include agricultural chemicals that are bound to soil particles as evidenced by findings of Foxet al.(2010, p. 8840). Foxet al.(2010, p. 8840) determined that turbidity of streams in grotto sculpin caves in Perry County was positively correlated with total chemical and DEA concentrations. Additionally, Gerken and Adams (2007, p. 76) noted that siltation was a major problem in grotto sculpin sites and postulated that silt likely reduced habitat available to this fish.

Excessive siltation in aquatic systems can be problematic for fish because it can change the overall structure of the habitat (Berkman and Rabeni 1986, pp. 291-292). Silt can fill voids in rock substrate that are integral components of habitat for reproduction and predator avoidance. The grotto sculpin occurs in habitats with some level of sediment deposition (Gerken 2007, pp. 16-17, 23-25). However, siltation beyond what occurred historically could limit the amount of suitable habitat available (Gerken 2007, pp. 27-28; Gerken and Adams 2007, p. 76), and the threshold of siltation that renders cave habitat unsuitable for grotto sculpin has not yet been determined.

Industrial Sand Mining—Industrial sand is also known as “silica,” “silica sand,” and “quartz sand,” and includes sands with high silicon dioxide content. Silica sand production in the United States was 29.3 million metric tons (Mt), an increase of 5.3 Mt from 2009 to 2010 (U.S. Geological Survey (USGS) 2012, p. 66.6). The Midwest leads the Nation in industrial sand and gravel production, accounting for 49 percent of the annual total (USGS 2012, p. 66.1). One end-use of silica sand is as a propping agent for hydraulic fracturing. Higher production of silica sand in 2010 was primarily attributable to an increasing demand for hydraulic fracturing sand because of continuing exploration and production of natural gas throughout the United States. Conventional natural gas sources have become less abundant, leading drilling companies to turn to deep natural gas and shale gas. Of the 29.3 Mt of silica sand sold or used in the United States, 12.1 Mt (41 percent) was used for hydraulic fracturing in the petroleum industry (USGS 2012, p. 66.10). As of 2010, the price per ton for industrial silica sand was $45.24 in the United States (USGS 2012, p. 66.11). In addition to new facilities, existing hydraulic fracturing sand operations increased production capacity to meet the surging demand for sand.

Mining for silica sand in Missouri occurs in the St. Peter Sandstone in Jefferson, Perry, and St. Louis Counties (USGS 2011, p. 27.2). The St. Peter Sandstone formation is directly adjacent to (to the west) the Joachim Dolomite formation that forms the karst habitat for the grotto sculpin in Perry County. The interface between these two formations generally comprises the western borders of the Central Perryville and Mystery-Rimstone karst areas. Four companies in Missouri produced 0.9 Mt of high-purity sand from the St. Peter Sandstone formation (USGS 2011, p. 27.2). The existing operation in Perry County lies 5.6 km (3.5 mi) northwest of Perryville and involves open pit mining on 101 ha (250 acres). This producer specializes in 40 to 70 and 70 to 140 size-grades that were used by the oil and gas well-servicing industry as a hydraulic fracture propping agent in shale formations (USGS 2010, p. 27.2).

Sand mining is typically accomplished using open pit or dredging methods with standard mining equipment and without the use of chemicals. Sand can be mined from outcrops or by removing overburden to reach subsurface deposits. Environmental impacts of sand mining are primarily limited to disturbance of the immediate area. The current operation in Perry County is partially within the Joachim Dolomite formation and at the western edge of the sinkhole plain with approximately four sinkholes occurring in the immediate vicinity. Erosion of soil and disturbed overburden could occur and increase the sediment loads in adjacent surface waters and cave streams via runoff. For example, a portion of the existing mining operation is within the Bois Brule watershed. Sediment-laden runoff could enter Blue Spring Branch, one of the surface streams occupied by the grotto sculpin. As described above, sedimentation can change the structure of grotto sculpin habitat and negatively impact reproduction and predator avoidance. Presence of the current facility, only 0.5 km (0.3 mi) and 1.6 km (1 mi) from the Central Perryville Karst and Crevice Cave recharge area, respectively, shows that such operations can and do occur in the Joachim Dolomite formation and immediately adjacent to grotto sculpin habitat. We currently are unaware of any plans for new facilities or expansions of current facilities. However, based on the presence of one existing operation, the occurrence of St. Peter Sandstone in Perry County, as well as recent growth of the hydraulic fracturing industry and associated increased demand for silica sand, it is likely that increased sand mining activity will occur in the future in areas where the grotto sculpin occurs. We consider sand mining to be a potentially significant threat to the species in the future.

Summary of Factor A

All of the recharge areas for caves occupied by the grotto sculpin are highly vulnerable and contain hazards from historical sinkhole dumps, agricultural practices without universal application of best management practices, ineffective private septic systems, and degraded runoff from roads. Hazardous waste facilities, outfalls for waste and storm water, and underground storage tanks are found in the recharge area for Crevice Cave that are not found in other parts of the species' range. Cave recharge areas in the Central Perryville Karst contain an average of 23 sinkholes per km2(58 per mi2), whereas those in the Mystery-Rimstone Karst contain an average of 11 per km2(27 per mi2). Water contamination from various sources of point and non-point source pollution poses a significant, ongoing threat to the grotto sculpin. Water flow in karst systems occurs by way of surfacefeatures, such as sinkholes and losing streams, as well as connectivity to the underlying aquifer. Sinkholes can funnel storm-runoff that carries contaminants directly into cave systems in a short period of time and severely degrades water quality.

At least half of the sinkholes in Perry County have been, or are currently used as, dump sites for anthropogenic waste including household chemicals, sewage, pesticide and herbicide containers, and animal carcasses. Cave streams in Perry County are contaminated with current-use and legacy-use pesticides that enter cave systems through storm runoff or via leaching in sinkholes. The majority of chemicals that have TWAs at levels above MDLs originated from agricultural pest management activities and included acetochlor, diethatyl-ethyl, atrazine, and desethylatrazine (DEA), pyrene, metolachlor, DEET, and pentachloroanisole. Atrazine has been the most frequently detected herbicide in ground and surface waters in Perry County. Even at concentrations below EPA criteria for protection of aquatic life, atrazine has been shown to reduce egg production and cause gonadal abnormalities in fish.

Potential contaminant problems with sinkholes are further exacerbated by the presence and continued installation of vertical drains across the agricultural landscape. This practice, meant to reduce erosion by facilitating drainage of surface or subsurface water, results in more land available to the farmer. As of 2010, the recharge areas for known and potential grotto sculpin habitat in the Central Perryville and Mystery-Rimstone karst areas contain an average of 2.4 vertical drains per km2(6.2 per mi2). Vertical drains have been linked directly to contamination of groundwater and water used for human consumption. Vertical drains also act as attractive nuisances because, like sinkholes, they are potential targets for illegal dumping of hazardous waste.

Risk from agricultural land use and point sources of pollution, such as sinkhole dumps, are not the only concern on the Perry County landscape. The recharge area for Crevice Cave contains the city of Perryville. Urban growth and hazards, such as hazardous waste facilities, underground storage tanks, wastewater discharges, and poorly maintained septic systems, in and around the city are threats to water quality in the range of the grotto sculpin. Potential threats in more rural areas of Perry County include introduction of manure and associated bacteria and pathogens into sinkholes from small livestock feeding areas. Such contaminants can increase nutrient loads and lower dissolved oxygen in the groundwater.

Concerns with sedimentation and wash load are primarily the transport of contaminants and the deposition of sediment in cave streams. Turbidity of cave streams is positively correlated with chemical concentrations, indicating that chemicals can bind to sediment particles and be transported by surface runoff. Siltation beyond what occurred historically could limit the amount of suitable habitat available; abnormally high deposition of sediment in cave systems can be problematic for aquatic life as it can fill voids in rock substrate that are integral components of grotto sculpin habitat.

Industrial sand mining is occurring in Perry County just outside the range of the grotto sculpin, but within the Bois Brule watershed. The mining operation near Perryville lies in the interface between the St. Peter Sandstone and Joachim Dolomite formations. Current mining operations could exacerbate erosion and sedimentation problems in the sinkhole plain and negatively impact grotto sculpin habitat. Furthermore, anticipated expansions of current operations or development of new operations to meet increasing demand of silica sand could pose a more serious threat in the future.

Factor B. Overutilization for Commercial, Recreational, Scientific, or Educational Purposes

Although some specimens of the grotto sculpin have been taken for scientific investigations, we do not consider such collection activities to be at a level that poses a threat to the species. We do not have records of any individuals being taken for commercial, recreational, or educational purposes.

Factor C. Disease or Predation

Predation by invasive, epigean fish poses a threat to eggs, young-of-year, and juvenile grotto sculpin. Farm ponds are human-made features, as opposed to natural aquatic habitats, that often are stocked with both native and nonnative fishes for recreational purposes. Fish from farm ponds enter cave systems through sinkholes when ponds are unexpectedly drained (Burret al.2001, p. 284) or after high-precipitation events. Predatory fish were documented to occur in all of the caves occupied by the grotto sculpin, and include common carp (Cyprinus carpio), fathead minnow (Pimephales promelas),yellow bullhead (Ameiurus natalis), green sunfish (Lepomis cyanellus), bluegill (Lepomis macrochirus), and channel catfish (Ictalurus punctatus) (Burret al.2001, p. 284).

The migration and persistence of invasive, epigean fish species into cave environments poses an ongoing and pervasive threat to the grotto sculpin because of unnatural levels of predation on eggs, young-of-year, and juveniles. Predation beyond what occurs naturally among adult and juvenile grotto sculpin can reduce population levels to an unsustainable level and may render a population unrecoverable in the face of an unexpected mass mortality.

Factor D. The Inadequacy of Existing Regulatory Mechanisms

The primary causes of the grotto sculpin's decline are degradation of aquatic resources from illegal waste disposal in sinkhole dumps, chemical leaching, urban development, and sedimentation. Existing Federal, State, and local laws have not been able to prevent impacts to the grotto sculpin and its habitat, and the existing regulatory mechanisms are not expected to prevent causes of grotto sculpin decline in the future.

The grotto sculpin is not protected under the Missouri State Endangered Species Law (MO ST 252.240) because it has not been formally recognized as a distinct species, but is affo