Cory O. Brant

A synthesized pheromone induces upstream movement in female sea lamprey and summons them into traps

Female insect pheromone blends induce robust tracking responses in males and direct them into traps. In vertebrates, pheromones that induce strong and precise tracking responses in natural habitats have rarely been described. Here, we show in the sea lamprey (Petromyzon marinus), a vertebrate invader of the Laurential Great Lakes, that a synthesized component of the male mating pheromone, 7α, 12α, 24-trihydroxy-5α-cholan-3-one 24-sulfate (3kPZS), when released into a stream to reach concentrations of 10−14, 10−13, 10−12, 10−11, or 10−10 M, triggers robust upstream movement in ovulated females drawing ≈50% into baited traps. Experiments conducted in diverse stream segments demonstrate the level of behavioral response was not affected by habitat conditions and is effective over hundreds of meters. 3kPZS is equally effective at luring ovulated females as the whole pheromone blend released by males between 10−14 and 10−11 M. 3kPZS diverts ovulated females away from and disrupts orientation to male washings when applied at concentrations higher than washings. Indeed, a single pheromone compound is able to redirect female sea lampreys away from a natural pheromone source and lure them into traps, which should be more effective than targeting males when applied in population control. Our findings may spur the discovery of other potent and environmentally benign agents to combat biological invasion, a process accelerated by globalization, exacerbated by climate change, and costing the global economy US

Characterization of a Novel Bile Alcohol Sulfate Released by Sexually Mature Male Sea Lamprey (Petromyzon marinus)

1.4 trillion of damage annually.

Quantification of a male sea lamprey pheromone in tributaries of laurentian great lakes by liquid chromatography-tandem mass spectrometry

A sulphate-conjugated bile alcohol, 3,12-diketo-4,6-petromyzonene-24-sulfate (DKPES), was identified using bioassay-guided fractionation from water conditioned with sexually mature male sea lamprey (Petromyzon marinus). The structure and relative stereochemistry of DKPES was established using spectroscopic data. The electro-olfactogram (EOG) response threshold of DKPES was 10−7 Molar (M) and that of 3-keto petromyzonol sulfate (3 KPZS; a known component of the male sea lamprey sex pheromone) was 10−10 M. Behavioural studies indicated that DKPES can be detected at low concentrations by attracting sexually mature females to nests when combined with 3 KPZS. Nests baited with a mixture of DKPES and 3 KPZS (ratio 1∶29.8) attracted equal numbers of sexually mature females compared to an adjacent nest baited with 3 KPZS alone. When DKPES and 3 KPZS mixtures were applied at ratios of 2∶29.8 and 10∶29.8, the proportion of sexually mature females that entered baited nests increased to 73% and 70%, respectively. None of the sexually mature females released were attracted to nests baited with DKPES alone. These results indicated that DKPES is a component of the sex pheromone released by sexually mature male sea lamprey, and is the second biologically active compound identified from this pheromone. DKPES represents the first example that a minor component of a vertebrate pheromone can be combined with a major component to elicit critical sexual behaviors. DKPES holds considerable promise for increasing the effectiveness of pheromone-baited trapping as a means of sea lamprey control in the Laurentian Great Lakes.

Chemical cues and pheromones in the sea lamprey (Petromyzon marinus)

We developed an assay for measuring 7α,12α,24-trihydroxy-5a-cholan-3-one-24-sulfate (3kPZS), a mating pheromone released by male sea lampreys (Petromyzon marinus), at low picomolar concentrations in natural waters to assess the presence of invasive populations. 3kPZS was extracted from streamwater at a rate of recovery up to 90% using a single cation-exchange and reversed-phase mixed-mode cartridge, along with [(2)H(5)]3kPZS as an internal standard, and quantified using ultrahigh performance liquid chromatography-tandem mass spectrometry. The limit of detection was below 0.1 ng L(-1) (210 fM), which was the lowest concentration tested. Intra- and interday coefficients of variation were between 0.3-11.6% and 4.8-9.8%, respectively, at 1 ng 3kPZS L(-1) and 5 ng 3kPZS L(-1). This assay was validated by repeat measurements of water samples from a stream spiked with synthesized 3kPZS to reach 4.74 ng L(-1) or 0.24 ng L(-1). We further verified the utility of this assay to detect spawning populations of lampreys; in the seven tributaries to the Laurentian Great Lakes sampled, 3kPZS concentrations were found to range between 0.15 and 2.85 ng L(-1) during the spawning season in known sea lamprey infested segments and were not detectable in uninfested segments. The 3kPZS assay may be useful for the integrated management of sea lamprey, an invasive species in the Great Lakes where pheromone-based control and assessment techniques are desired.

(+)- and (−)-Petromyroxols: Antipodal Tetrahydrofurandiols from Larval Sea Lamprey (Petromyzon marinus L.) That Elicit Enantioselective Olfactory Responses

Chemical cues and pheromones guide decisions in organisms throughout the animal kingdom. The neurobiology, function, and evolution of olfaction are particularly well described in insects, and resulting concepts have driven novel approaches to pest control. However, aside from several exceptions, the olfactory biology of vertebrates remains poorly understood. One exception is the sea lamprey (Petromyzon marinus), which relies heavily upon olfaction during reproduction. Here, we provide a broad review of the chemical cues and pheromones used by the sea lamprey during reproduction, including overviews of the sea lamprey olfactory system, chemical cues and pheromones, and potential applications to population management. The critical role of olfaction in mediating the sea lamprey life cycle is evident by a well-developed olfactory system. Sea lamprey use chemical cues and pheromones to identify productive spawning habitat, coordinate spawning behaviors, and avoid risk. Manipulation of olfactory biology offers opportunities for management of populations in the Laurentian Great Lakes, where the sea lamprey is a destructive invader. We suggest that the sea lamprey is a broadly useful organism with which to study vertebrate olfaction because of its simple but well-developed olfactory organ, the dominant role of olfaction in guiding behaviors during reproduction, and the direct implications for vertebrate pest management.

Petromyzonin, a hexahydrophenanthrene sulfate isolated from the larval sea lamprey (Petromyzon marinus L.).

(+)- and (-)-petromyroxol [(+)-1 and (-)-1, respectively], two novel tetrahydrofuran (THF)-diol fatty acid enantiomers, were isolated from water conditioned with larval sea lamprey. We herein describe their isolation and subsequent resolution using chiral chromatography. The absolute configuration of each enantiomer was determined by a combination of Mosher ester analysis and comparison with related natural and synthetic products. Electro-olfactogram (EOG) assays indicated that (+)-petromyroxol (1) possesses potent olfactory activity for sea lamprey.

Biosynthesis and release of pheromonal bile salts in mature male sea lamprey

A new hexahydrophenanthrene sulfate was identified from water conditioned with sea lamprey larvae ( Petromyzon marinus ) and named petromyzonin. Its structure was unequivocally elucidated on the basis of spectroscopic analyses including comparison with spectra of known compounds. The absolute configuration was determined by electronic circular dichroism. Petromyzonin may function as a chemical signal, as it elicited responses in electro-olfactogram recording with a dynamic concentration-response relationship and a detection threshold of 10(-11) M.

Isolation and identification of petromyzestrosterol, a polyhydroxysteroid from sexually mature male sea lamprey (Petromyzon marinus L.)

BackgroundIn vertebrates, bile salts are primarily synthesized in the liver and secreted into the intestine where they aid in absorption of dietary fats. Small amounts of bile salts that are not reabsorbed into enterohepatic circulation are excreted with waste. In sexually mature male sea lamprey (Petromyzon marinus L.) a bile salt is released in large amounts across gill epithelia into water where it functions as a pheromone. We postulate that the release of this pheromone is associated with a dramatic increase in its biosynthesis and transport to the gills upon sexual maturation.ResultsWe show an 8000-fold increase in transcription of cyp7a1, a three-fold increase in transcription of cyp27a1, and a six-fold increase in transcription of cyp8b1 in the liver of mature male sea lamprey over immature male adults. LC–MS/MS data on tissue-specific distribution and release rates of bile salts from mature males show a high concentration of petromyzonol sulfate (PZS) in the liver and gills of mature males. 3-keto petromyzonol sulfate (3kPZS, known as a male sex pheromone) is the primary compound released from gills, suggesting a conversion of PZS to 3kPZS in the gill epithelium. The PZS to 3kPZS conversion is supported by greater expression of hsd3b7 in gill epithelium. High expression of sult2b1 and sult2a1 in gill epithelia of mature males, and tissue-specific expression of bile salt transporters such as bsep, slc10a1, and slc10a2, suggest additional sulfation and transport of bile salts that are dependent upon maturation state.ConclusionsThis report presents a rare example where specific genes associated with biosynthesis and release of a sexual pheromone are dramatically upregulated upon sexual maturation in a vertebrate. We provide a well characterized example of a complex mechanism of bile salt biosynthesis and excretion that has likely evolved for an additional function of bile salts as a mating pheromone.

Multiplex quantification of lamprey specific bile acid derivatives in environmental water using UHPLC-MS/MS.

Petromyzestrosterol (1), a novel polyhydroxylated steroid, was identified from water conditioned with sexually mature male sea lamprey (Petromyzon marinus), a jawless vertebrate animal. Along with this novel steroid, two known steroids, 7α,12α,24-trihydroxy-5α-cholan-3-one-24-sulfate (3k PZS) and 7α,12α,24-trihydroxy-5α-cholan-24-sulfate (PZS), were isolated. Structures of these compounds were unequivocally established by spectroscopic analyses and by comparison with spectra of known compounds. Electro-olfactogram recordings (EOG) showed that 1 at nanomolar concentrations was stimulatory for the olfactory epithelium of adult females. 3k PZS, known to function as a male sex pheromone, was more stimulatory than 1 for the female olfactory epithelia. The concentration-response curve of 3k PZS was exponential in shape with steep slopes between 10(-10) and 10(-6) mol L(-1). The concentration-response curve for 1 was shallower than that for 3k PZS.

A thermogenic secondary sexual character in male sea lamprey

Larval and adult sea lampreys (Petromyzon marinus) release bile salts and acids into the surrounding aquatic environment. Some of these bile salts and acids, such as petromyzonol sulfate (PZS), 3-keto petromyzonol sulfate (3k PZS), petromyzonamine disulfate (PADS), petromyzosterol disulfate (PSDS), and 3-keto allocholic acid (3k ACA), may function as pheromones. To examine the release and distribution patterns of these metabolites, which this study has termed bile acid derivatives, we developed a novel UHPLC-MS/MS method that was characterized by simple sample preparation, baseline separation, and short analysis time for all studied compounds. These five analytes were separated in 7 min using a reversed-phase C18 column containing 1.7 μm particles and a gradient elution at pH 8.9. Once separated, the analytes were subjected to electrospray ionization-mass spectrometry (negative ion mode) and collision-induced dissociation tandem mass spectrometry (CID-MS/MS) using the multiple reaction monitoring (MRM) mode. Deuterated 3k PZS ([(2)H(5)]3k PZS) was added as the internal standard (IS) to the sample prior to solid phase extraction (SPE). Among the three types of SPE sorbent tested, mixed-mode cation-exchange and reversed-phase sorbent for bases (MAX) and acids (MCX), and reversed-phase C18 sorbent (Sep-pak), the best recoveries (84.1-99.7%) were obtained with MCX cartridges. The calibration curves of all five analytes were linear between 0.15 and 1200 ng/mL, with R(2)≥0.9997. This method had a precision of relative standard deviation (RSD) ≤9.9% and an accuracy of deviation (DEV) ≥92.5%. The developed method was successfully used to quantify bile acid derivatives found in streams where lampreys spawn (SD<1.4) and water conditioned with male sea lampreys (SD<4.8). Utilizing this method provides a routine analysis of lamprey bile acid derivatives and may prove useful for sea lamprey population estimates in future studies and applications.