Kontinuirani monitoring stanja životne sredine: Optimizacija sistema zasnovanog na fiziološkim odgovorima dagnje (Mytilus galloprovincialis Lamarck, 1819)

Abstract

U okviru ove disertacije optimizovan je sistem za kontinuirano praćenje statusa voda obalnog mora zasnovan na fiziološkim biomarkerima mediteranske dagnje – srčanog ritma i pokreta ljušture. Ispitana je pouzdanost sistema za rano upozorenje, varijabilnostfizioloških biomarkera kod dagnje, optimalno vrijeme korišćenja odabranog bioindikatora u procesu kontinuiranog biomonitoringa, kao i nedostaci postojećeg biomonitoring sistema u cilju njegovog daljeg usavršavanja i implementacije, uključujući i unapređenja postojećeg automatizovanog sistema za praćenje srčane aktivnosti u dijelu vezanom za transfer i obradu signala. Rezultati praćenja bazalnog srčanog ritma i pokreta ljuštura dagnji u kontrolnim uslovima, bez prisustva stresora, su pokazali kako individualnu, tako i grupnu varijabilnost ovih biomarkera kod jedinki u monitoring sistemu u funkciji vremena.Međutim, bez obzira na individualne varijacije, eksponirane jedinke su pokazale isti trend ponašanja, pa se mogu posmatrati kao grupa. Nije pokazan izražen cirkadijalni ni sezonski karakter odabranih fizioloških biomarkera. Ispitana je osjetljivost odabrane vrste i automatizovanog ex situ sistema na prirodna variranja ekoloških faktora praćenih mjesečnom dinamikom. Pokazalo se da srčani ritam ne zavisi od variranja ekoloških faktora u rangu uobičajenih vrijednosti karakterističnih za odabrani lokalitet. ukupna kondicija jedinki, a time i performanse monitoring sistema provjerene su periodičnim izlaganjem sistema kratkotrajnom ekstremnom stresu – naglom smanjenju saliniteta. Zbirni odgovor grupe jedinki u monitoring sistemu na stres praćen je mjesečnom dinamikom. Odabrani bomarkeri – pokreti ljušture, varijabilnost srčanog ritma i vrijeme potrebno za oporavak srčanog ritma nakon kratkotrajnog izlaganja stresu su se pokazali kao dobri indikatori kondicionog statusa jedinki. Rezultati testova saliniteta su pokazali da su pokreti ljušture robusniji biomarker od srčanog ritma, jer je konzistentan odgovor grupe bilježen u svim testovima saliniteta tokom jednogodišnjeg monitoringa. Konstantno produžavanje perioda potrebnog za oporavak srčanog ritma nakon prestanka dejstva stresora već od drugog mjeseca monitoringa, kao i potpuni izostanak reakcije u smislu promjena srčanog ritma usled djelovanja stresora tokom poslednja četiri mjeseca monitoringa su pokazali da jedinke u monitoring sistemu imaju ograničeni optimalni vijek. Utvrđeno je da već nakon tri do četiri mjeseca kontinuiranog monitoringa dolazi do “zamora” sistema. Optimalno vrijeme korišćenja odabranog bioindikatora iznosi tri mjeseca. Jedinke u sistemu je potrebno zamijeniti na svakih tri mjeseca, kako bi se povećala pouzdanost sistema za rano upozorenje. U cilju unapređenja postojećeg automatizovanog sistema za praćenje srčanog ritma realizovano je novo rješenje u domenu prenosa podataka sa mjerne opreme na računar. Bežična razmjena podataka ne degradira performanse cjelokupnog sistema. Primjenjivost odabranih fizioloških biomarkera u pasivnom biomonitoringu na nativnim jedinkama mediteranske dagnje sa odabranih lokaliteta u gradijentu zagađenja ispitana je na odabranoj studiji slučaja u Bokokotorskom zalivu. Rezultati su pokazali da se metodologija zasnovana na koeficijentu varijacije i vremenu oporavka srčanog ritma nakon kratkotrajnog stresa u principu može koristiti u faznom pasivnom biomonitoringu na lokalitetima u gradijentu zagađenja. Da bi rezultati dobijeni praćenjem promjena srčanog ritma nakon izlaganja kratkotrajnom stresu jednoznačno ukazali na problem zagađenja na odabranim lokalitetima, stresor ne bi smio biti iz grupe abiotičkih faktora na koje se različite populacije odabrane vrste lako adaptiraju.

The focus of this thesis was the optimization of ex situ costal waters biomonitoring system based on two physiological biomarkers of Mediterranean mussel, namely heart rate and valve gape. Experimental set-up provided not only insight into reliability of the early warning system and variability of physiological biomarkers over a year-long continuous operation, but enabled defining the optimal shelf life of bioindicator organism in the continuous biomonitoring. Some shortcomings of the existing automated system formonitoring cardiac activity as well as steps for further improvement and implementation were identified, including improvements related to signal transfer and processing. Basal heart rate and valve gape dynamics in control conditions, with no significant stress,demonstrated considerable time – dependant individual and group variability. Still, the responses of all tested specimens, regardless of their individual variability, followed a similar behavioural trend, which implies that the test individuals can be clearly regarded as a group. The responses of selected physiological biomarkers did not follow circadian or seasonal pattern. The sensitivity of automated ex situ system and selected species to natural variation of a number of environmental variables has also been investigated. The results indicate that the mussels’ heart rate does not depend on key environmental factors provided they remain within the usual variation range typical for the selected site. The overall physiological condition of test individuals and performances of monitoring system have been tested on monthly basis by group response to short-term exposure to extreme stress conditions (a sharp salinity decrease). Selected biomarkers, valve gape, coefficient of variation and recovery time of heart rate after the exposure to stress, proved to be good indicators of condition of individuals in monitoring system. Valve gape turned out as a more robust biomarker compared to the heart rate, as consistent group response was recorded in all salinity tests throughout the experiment. Successive increase of the heart rate recovery time was recorded since the second month of monitoring till the end of the monitoring, while the complete absence of an expected reaction to stress – change of the heart rhythm – was apparent during the last four months of monitoring. These results indicate that the individuals in the monitoring system have limited optimal shelf life and that after three to four months of continuous monitoring, the system is subject to "fatigue". It can be concluded that the optimal shelf life of the individual mussels in biomonitoring system is three months. To maintain the reliability of the early warning system it is advisable to replace individuals in the system every three months. In order to improve the existing automated system for heart rate monitoring, a new solution for data transfer from the sensor to the computer was implemented. Wireless data exchange did not degrade the performance of the entire system. A case study in Boka Kotorska Bay was used to assess the applicability of selected physiological biomarkers in passive biomonitoring usingnative Mediterranean mussel specimens from selected locations in pollution gradient. The results showed that the methodology based on the coefficient of variation and heart rate recovery time after short-term exposure to extreme stress can, in general, be used in periodical passive biomonitoring of sites in the pollution gradient. However, to diminish the impact of confounding factors, to increase the credibility of the environmental assessment of selected polluted sites based on hart rate changes after exposure to shortterm stress, the stressor should not be chosen from the group of abiotic factors to which different populations of the selected species easily adapt.