This protocol describes the use of biomarkers for the early detection of deleterious impacts in aquatic ecosystems. The biomarkers are closely related to sentinel traits, and their changes aid in detecting early-warning damages.
The larvae of dobsonflies from the genus Corydalus, commonly known as Hellgrammites, are characterized by their notable size, extensive range of occurrence, and extended period of immaturity, which can last up to one year. Hellgrammites are clearly known to exhibit sensitivity to pollution and habitat structure impacts. Given these unique features, using Corydalus texanus larvae is highly suitable as reliable biomonitoring agents to assess the ecological integrity of aquatic ecosystems. This protocol aims to provide the necessary tools for C. texanus assessment and demonstrate their efficacy through a case study. Research findings have practical implications, indicating that C. texanus larvae exhibit early-warning responses to mining pollution, bioaccumulating high amounts of heavy metals such as Zn, Fe, and Al. The presence or absence of C. texanus populations may serve as a helpful indicator for identifying potential issues related to ecosystem health. The unconventional approach has shown early warnings of pollution in mining-impacted sites, highlighting the need for timely action to protect the environment. Given their unique traits, the use of C. texanus larvae is highly suggested as a reliable non-conventional bioindicator.
Hellgramites are insect larvae from the order Megaloptera (Latreille, 1802), named dobsonflies or fishflies in their adult stage. A low diversity but widespread characterizes this group of top predator insect larvae from aquatic ecosystems1. Hellgramite species occur in well-defined biogeographic regions; therefore, it is relatively easy to identify species without a high taxonomic knowledge. Notably, the Corydalidae larvae possess the most prominent species from the Megaloptera order (20-90 mm body length)2, making hellgramite visible to the naked eye.
Hellgrammites play a crucial role in aquatic ecosystems as predators, with a powerful presence due to large chews denoting their impressive predatory shape. A dorsoventrally flattened body also joins with 7-8 pairs of filament gills along the body, and a head capsule with six stemmata per side makes hellgrammites fascinating organisms for entomologists and fans3. Adults of Corydalidae surprise and create an impression image to people due to their prominent size; however, they are entirely harmless. It is noteworthy that hellgrammites have the ability to persist in aquatic environments in their larval stage for a significant duration.
The phenotypical features of hellgrammites allow a particular chance to highlight their role in aquatic ecosystems; nevertheless, their indicator potential is the most wanted feature for aquatic ecologists. Vast knowledge of their bioindicator potential is highlighted in aquatic ecosystems because their occurrence relates to good health conditions in their habitats due to their intolerance to organic pollution in surface waters4,5,6,7,8.
Most Corydalidae megalopterans live in high-speed running waters as riffles and substrates predominated by cobble and pebbles, but hellgrammites also occur in low-gradient streams with snags and sand substrates, as well as in lentic habitats such as lakes3,9,10. Their wide range of occurrence mirrors the critical traits of a top predator and their ability to colonize several habitats aimed by their effective life history strategies11. Hellgramites linked their traits with the dynamics of aquatic ecosystems; thus, strategies such as adaptation to aerial respiration by spiracles (in addition to their ventral tufts of tracheal gills) are proper of the Corydalidae strategies10.
Hellgrammites inhabit particular ecosystems and exhibit rapid responses to deviations from established patterns, thereby serving as an early-warning system7. The reactions of these wildlife organisms can be employed as a valuable tool to assess the impact of pollution on aquatic ecosystems, particularly in the case of non-target pollutant mixtures. Some deleterious effects on living organisms have been recognized in ecosystems because of individual toxicity of chemicals, but the effect of pollutant mixtures shall be identified. The early-warning responses from hellgrammites may make it possible to identify deleterious effects by giving a reference to the impacts of a mixture of pollutants or even when individual effects of pollutants recognize No Observable Effect Concentration (NOEC)12.
Several model organisms have been used for experimental acute and chronic tests; however, they are cultured and maintained under controlled conditions13. Controlled conditions make them unable to identify the non-target effects of several pollutants they are exposed to. Also, NOEC is frequently recognized due to the complexity of the pollutant's mixture. For that reason, in the last decades, non-model native species with non-target effects were recognized to screen systems, which are essential to carrying out ecotoxicological research14. Consequently, hellgrammites seem able to assess the deleterious effects of pollution in aquatic ecosystems. Traits such as their biology, genetics, and physiology, among others, make non-model organisms suitable for impact assessment in ecosystems14.
This protocol aims to establish a novel biomonitoring tool using a non-model organism that can detect early-warning signals in response to non-target pollution mixtures. To achieve the best results, the traits exhibited by the larvae of the model species, C. texanus, have been comprehensively considered and integrated into the analysis15.
Although the use of C. texanus is optimal for evaluation, it is necessary to consider several aspects of its use and collection. Chosen study sites prove challenging, owing to several factors such as unfavorable weather conditions, geographical inaccessibility, high aridity levels, or insufficient security protocols in selected regions. Stated constraints and limitations can often present challenges when conducting fieldwork. When evaluating specific regions, particularly those in tropical streams, obtaining sam…
The authors have nothing to disclose.
The authors would like to express their sincere gratitude to CONAHCyT for providing the FONINS P 1931 grant, which greatly facilitated their research efforts. They would also thank the Secretaría de Investigación y Posgrado at Instituto Politécnico Nacional for the invaluable support provided through the SIP project grant (20200577). In addition, the first author wishes to acknowledge the generous post-graduate scholarship awarded by CONAHCyT, which enabled the team to conduct field trips and gather essential data. Finally, the authors would like to express their appreciation for the invaluable assistance of María Teresa García Camacho in the laboratory, without whom this project would not have been possible.
Analytical balance of 220 g | Ohaus | PR224/E | This balance is useful for weighing the extracted tissue from specimens. |
Chest waders | LaCrosse | 700152M | We recommend using waders for sample collection. Alternatively, you can also use boots with rubber hip boots for sampling. |
Cutting board | True | TRUE915121 | It is recommended to use white and plastic boards. |
Forceps | DR Instruments | 112 | It is recommended to use a 12 pul forceps made of stainless steel. |
Inductively Coupled Plasma-Optical Emission Spectroscopy System | Perkin Elmer | 7300 DV | A device for quantifying heavy metals using spectroscopy. |
Kick net | LaMotte | 0021-P | An alternative method for making a kick net involves manually crafting one using a mesh with a thickness of 500 micrometers. |
Liquid Nitrogen | NA | NA | NA |
Liquid Nitrogen Dewar Static Cryogenic Container | BestEquip | DF0504 | It is recommended to use an aluminum tank with canisters. |
Mortar and Pestle Set | Cole-Parmer | EW-63100-54 | This is a porcelain mortar and pestle used for grinding dry tissues. |
Multiwave GO Plus | Anton Paar | C93IP001EN-E | Multiwave is a useful tool for digesting multiple samples. |
Oven for drying | Fisher scientific | 506G | An incubator oven, also known as a dry tissue oven, is essential for drying tissues at temperatures of at least 80°C. |
Precision scalpel | Xcelite by Weller | 037103-48768 | It is recommended to use a scalpel made of aluminum. |
Tissue Homogenizer (tearer) | Kopro | K110000 | It is recommended to use a tissue tearer with a base. Some companies offer ultrasonic tearers, which may be the optimal choice. |
Ultra-Low Temperature Chest Freezer | REVCO | CA89200-384 | Many companies provide freezers, but it is recommended to choose one with a storage temperature of at least -40°C. |
Wide mouth plastic bottles | United Scientific | 81900 | Using polypropylene bottles with wide caps and mouths is strongly recommended. |
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