Jon J. Amberg

Fish gut microbiota analysis differentiates physiology and behavior of invasive Asian carp and indigenous American fish

Gut microbiota of invasive Asian silver carp (SVCP) and indigenous planktivorous gizzard shad (GZSD) in Mississippi river basin were compared using 16S rRNA gene pyrosequencing. Analysis of more than 440 000 quality-filtered sequences obtained from the foregut and hindgut of GZSD and SVCP revealed high microbial diversity in these samples. GZSD hindgut (GZSD_H) samples (n=23) with >7000 operational taxonomy units (OTUs) exhibited the highest alpha-diversity indices followed by SVCP foregut (n=15), GZSD foregut (n=9) and SVCP hindgut (SVCP_H) (n=24). UniFrac distance-based non-metric multidimensional scaling (NMDS) analysis showed that the microbiota of GZSD_H and SVCP_H were clearly separated into two clusters: samples in the GZSD cluster were observed to vary by sampling location and samples in the SVCP cluster by sampling date. NMDS further revealed distinct microbial community between foregut to hindgut for individual GZSD and SVCP. Cyanobacteria, Proteobacteria, Actinobacteria and Bacteroidetes were detected as the predominant phyla regardless of fish or gut type. The high abundance of Cyanobacteria observed was possibly supported by their role as the fish’s major food source. Furthermore, unique and shared OTUs and OTUs in each gut type were identified, three OTUs from the order Bacteroidales, the genus Bacillariophyta and the genus Clostridium were found significantly more abundant in GZSD_H (14.9–22.8%) than in SVCP_H (0.13–4.1%) samples. These differences were presumably caused by the differences in the type of food sources including bacteria ingested, the gut morphology and digestion, and the physiological behavior between GZSD and SVCP.

Persistence of DNA in carcasses, slime and avian feces may affect interpretation of environmental DNA data

The prevention of non-indigenous aquatic invasive species spreading into new areas is a goal of many resource managers. New techniques have been developed to survey for species that are difficult to capture with conventional gears that involve the detection of their DNA in water samples (eDNA). This technique is currently used to track the invasion of bigheaded carps (silver carp and bighead carp; Hypophthalmichthys molitrix and H. nobilis) in the Chicago Area Waterway System and Upper Mississippi River. In both systems DNA has been detected from silver carp without the capture of a live fish, which has led to some uncertainty about the source of the DNA. The potential contribution to eDNA by vectors and fomites has not been explored. Because barges move from areas with a high abundance of bigheaded carps to areas monitored for the potential presence of silver carp, we used juvenile silver carp to simulate the barge transport of dead bigheaded carp carcasses, slime residue, and predator feces to determine the potential of these sources to supply DNA to uninhabited waters where it could be detected and misinterpreted as indicative of the presence of live bigheaded carp. Our results indicate that all three vectors are feasible sources of detectable eDNA for at least one month after their deposition. This suggests that current monitoring programs must consider alternative vectors of DNA in the environment and consider alternative strategies to minimize the detection of DNA not directly released from live bigheaded carps.

Sexually dimorphic gene expression in the gonad and liver of shovelnose sturgeon (Scaphirhynchus platorynchus)

The development of a biomarker for rapid detection of intersex is desirable to researchers and fisheries managers alike. The first step is to identify a marker that has a dimorphic expression pattern. With a focus on sex-specific genes, we assessed the use of vitellogenin (vtg), forkhead box L2 (foxl2) and doublesex and mab-3-related transcription factor 1 (dmrt1) as molecular biomarkers for the identification of gender in shovelnose sturgeon (Scaphirhynchus platorynchus), a species known to have intersex individuals. A total of 61 shovelnose sturgeons were sampled from the Wabash River, Indiana, in April 2008. Results from our study indicate that least 7.5% of the non-females had testicular oocytes. Expression level of liver vtg was not higher in females than males, nor was gonad dmrt1 expression found to be higher in males. Only the expression of foxl2 was significantly greater in females and was found to be useful for identifying gender. Variation in expression levels of foxl2 in gonads of intersex fish limited its usefulness as a single biomarker for identifying this condition. Instead, the use of foxl2 to dmrt1 (foxl2 transcript abundance/dmrt1 transcript abundance) may be useful in the identification of intersex fish, as this ratio increased with increased feminization. We conclude that foxl2 can be used to identify sex, but not intersex sturgeon.

Carbon Dioxide as a Tool to Deter the Movement of Invasive Bigheaded Carps

AbstractNonnative bigheaded carps are established in the Mississippi River and there is substantial concern about their potential entry into the interconnected Laurentian Great Lakes. While electrical barriers currently exist as a preventative measure, there is need for additional control mechanisms to promote barrier security through redundancy. We tested the effectiveness of infused carbon dioxide gas (CO2) as a tool to influence the movement and behavior invasive bigheaded carps, namely Bighead Carp Hypophthalmichthys nobilis and Silver Carp H. molitrix, as well as native Bigmouth Buffalo Ictiobus cyprinellus, Channel Catfish Ictalurus punctatus, Paddlefish Polyodon spathula, and Yellow Perch Perca flavescens in an experimental pond. Individuals were monitored with acoustic telemetry before, during, and after CO2 addition to the pond. We noted distinct changes in fish behavior following CO2 addition. Each species except Paddlefish maintained farther distances from the CO2 infusion manifold relative to ...

Detecting the movement and spawning activity of bigheaded carps with environmental DNA

Bigheaded carps are invasive fishes threatening to invade the Great Lakes basin and establish spawning populations, and have been monitored using environmental DNA (eDNA). Not only does eDNA hold potential for detecting the presence of species, but may also allow for quantitative comparisons like relative abundance of species across time or space. We examined the relationships among bigheaded carp movement, hydrography, spawning and eDNA on the Wabash River, IN, USA. We found positive relationships between eDNA and movement and eDNA and hydrography. We did not find a relationship between eDNA and spawning activity in the form of drifting eggs. Our first finding demonstrates how eDNA may be used to monitor species abundance, whereas our second finding illustrates the need for additional research into eDNA methodologies. Current applications of eDNA are widespread, but the relatively new technology requires further refinement.

Sex-Specific Gonadal and Gene Expression Changes throughout Development in Fathead Minnow

Although fathead minnows (Pimephales promelas) are commonly used as a model fish in endocrine disruption studies, past studies have not characterized sex-specific baseline expression of genes involved in sex differentiation during development in this species. Using a sex-linked DNA marker to verify gender, we evaluated the expression over time of genes involved in sex differentiation (dmrt1, cyp19a, cyp17, star, esr1, ar) in developing fathead minnows (10-45 days post hatch). Evaluation of these molecular markers in combination with gender identification help us to better understand the mechanisms regulating sex differentiation in fathead minnows and how endocrine-disrupting chemicals may alter these processes.

Antagonists to the Wnt Cascade Exhibit Sex-Specific Expression in Gonads of Sexually Mature Shovelnose Sturgeon

Little is known regarding molecular mechanisms involved in sex determination and differentiation in sturgeon species. We addressed this knowledge gap by using next generation pyrosequencing technology to provide transcript libraries and species-specific sequences for mature gonads of shovelnose sturgeon, Scaphirhynchus platorynchus. We then mined these libraries to identify gender-specific transcripts and quantified relative transcript abundance using quantitative real-time polymerase chain reaction (qPCR). We detected a limited number of genes known to play a role in sex differentiation in other species. The sequence for dmrt1 was found only in the testes library. The abundance of dmrt1 differed slightly between the sexes, but the melt curve suggests that there may be different isoforms of dmrt1 in ovaries and testes of shovelnose sturgeon. The transcription factor foxl2 was 5.3-fold greater in ovaries than in testes. Two antagonists to the Wnt cascade, dickkopf-1 (dkk1) and dapper-1 (dact1), were found only in the ovary library. Results from qPCR indicated that dkk1 and dact1 were upregulated 1,819.1- and 207.5-fold, respectively, in ovaries compared with testes. These results suggest that antagonists to the Wnt cascade may play significant roles in sex differentiation and gonadal development in sturgeon and could serve as sex markers in this group of ancient fish.

Blood Chemistry Values for Shovelnose and Lake Sturgeon

Blood chemistry panels are commonly used for assessing the general health of vertebrate animals. Here, we present novel blood chemistry data for two North American sturgeon species, shovelnose sturgeon Scaphirhynchus platorynchus and lake sturgeon Acipenser fulvescens. Measurements were done using a portable chemistry analyzer (VetScan Analyzer; Abaxis). Among the plasma values measured (mean ± SD for shovelnose and lake sturgeon, respectively) were total proteins (3.7 ± 0.9 and 2.8 ± 0.4 g/dL), albumin (2.0 ± 0.5 and 1.1 ± 0.2 g/dL), globulin (1.7 ± 0.7 and 1.7 ± 0.3 g/dL), glucose (107 ± 46 and 62 ± 9.7 mg/dL), sodium (Na(+); 132 ± 3.6 and 150 ± 14 mEq/L), potassium (K(+); 3.5 ± 0.2 and 2.8 ± 1.7 mEq/L), phosphorus (10.4 ± 1.9 and 11.6 ± 3.6 mg/dL), and aspartate aminotransferase (AST; 676 ± 433 and 634 ± 234 IU/L). Higher values for total proteins, albumin, glucose, and Na(+) in shovelnose sturgeon than in lake sturgeon probably are the result of handling stress. In addition, the plasma of male shovelnose sturgeon had higher concentrations of AST, glucose, and globulin than did that of females, whereas the plasma of females had higher concentrations of albumin and K(+) than that of males. This study is the first to report blood chemistry data for shovelnose sturgeon. Robust blood chemistry databases can be used by aquaculturists and fish managers for monitoring sturgeon health.

Comparing Efficiency of American Fisheries Society Standard Snorkeling Techniques to Environmental DNA Sampling Techniques

AbstractAnalysis of environmental DNA (eDNA) is an emerging technique used to detect aquatic species through water sampling and the extraction of biological material for amplification. Our study compared the efficacy of eDNA methodology to American Fisheries Society (AFS) standard snorkeling surveys with regard to detecting the presence of rare fish species. Knowing which method is more efficient at detecting target species will help managers to determine the best way to sample when both traditional sampling methods and eDNA sampling are available. Our study site included three Navajo Nation streams that contained Navajo Nation Genetic Subunit Bluehead Suckers Catostomus discobolus and Zuni Bluehead Suckers C. discobolus yarrowi. We first divided the entire wetted area of streams into consecutive 100-m reaches and then systematically selected 10 reaches/stream for snorkel and eDNA surveys. Surface water samples were taken in 10-m sections within each 100-m reach, while fish presence was noted via snorkeli...

Comparison of American Fisheries Society (AFS) Standard Fish Sampling Techniques and Environmental DNA for Characterizing Fish Communities in a Large Reservoir

AbstractRecently, methods involving examination of environmental DNA (eDNA) have shown promise for characterizing fish species presence and distribution in waterbodies. We evaluated the use of eDNA for standard fish monitoring surveys in a large reservoir. Specifically, we compared the presence, relative abundance, biomass, and relative percent composition of Largemouth Bass Micropterus salmoides and Gizzard Shad Dorosoma cepedianum measured through eDNA methods and established American Fisheries Society standard sampling methods for Theodore Roosevelt Lake, Arizona. Catches at electrofishing and gillnetting sites were compared with eDNA water samples at sites, within spatial strata, and over the entire reservoir. Gizzard Shad were detected at a higher percentage of sites with eDNA methods than with boat electrofishing in both spring and fall. In contrast, spring and fall gillnetting detected Gizzard Shad at more sites than eDNA. Boat electrofishing and gillnetting detected Largemouth Bass at more sites t...