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Business potential in developing selective forms of tourism is found in all areas of Croatia, and development of health tourism is essential to the future of this country. Health tourism reflects its future through development of wellness offers, treatment, as well as medical tourism. Physical and rehabilitation medicine, through the domain of promoting health and preventing illnesses, along with a crucial rehabilitation aspect, reaches its full potential in the sector of health tourism, specifically continuous multidisciplinary collaboration of health and other professionals. Recognising capacity of continental part of Croatia as part of health tourism is only possible via continuous multidisciplinary cooperation, as well as enhancing necessary competencies and skill of all new and existing businesspersons. Business development within health tourism is possible via continuous promotion on a national, as well as international market with a goal of developing international recognisability and concurrency, observing new technological enhancements and educational advances in this sector. Sustainable and stable growth of health tourism is the ultimate goal which will result in opening new work positions, retaining younger population in the domicile and employment. Covid-19 pandemic resulted in developing new programs of rehabilitation within health tourism and enhancing existing programs in all regions of Croatia which is an important step towards the future this profession deserves.

Aluminium capsules can be used as the final packaging for any type of powdered foods, such as tea or vitamin and similar hot drinks. Aluminium represents a 100% barrier, is impermeable, extends the shelf life of the final product, is recyclable and an alternative to plastic. A wide range of individualized products, including the capsule as attractive single-use possibility, made it possible to satisfy the preferences and expectations of consumers. Currently two main materials for single serve portion coffee capsules are in use: aluminium and/or plastic based.

A multixenobiotic resistance (MXR) mechanism mediated by ABC binding cassette (ABC) transport proteins is an efficient chemical defence mechanism in sea urchin embryos. The aim of our work was to evidence whether exposure to sub- lethal doses of specific contaminants (oxybenzone (OXI), mercuric chloride (HgCl2), trybutiltin (TBT)) would induce MXR transporter activity during embryonic development (from zygote to blastula stage) in purple sea urchin (Paracentrotus lividus) embryos. Further, we present data on molecular identification, transport function, expression levels and gene localisation of two ABC efflux transporters – P- glycoprotein (ABCB1/P-gp) and sulfonylurea receptor like protein (ABCC9/SUR-like). Partial cDNA sequences of abcb1 and abcc9 were identified and quantitative PCR (qPCR) evidenced an increase in mRNA transcript levels of both ABC transporters during the 2-cell, as well as an overall decrease during the blastulae stage. Calcein- AM efflux activity assay indicated the activation of multidrug resistance-associated protein (MRP)/ABCC-like transport in the presence of HgCl2 and TBT in exposed blastulae. The in situ hybridisation of the 2-cell and blastula stages showed ubiquitous localisation of both transcripts within cells, supporting qPCR data. In conclusion, ABCB1 and ABCC9 are constitutive, as are HgCl2, TBT and OXI- inducible ABC membrane transporters, coexpressed in the zygote, 2-cell and blastula stages of the P. lividus. Their ubiquitous cell localisation further fortifies their protective role in early embryonic development.

Bisphenol A (BPA) presents a serious threat to soil ecosystems, yet its effects on soil- inhabiting organisms are mostly unexplored. Therefore, the impact of environmentally relevant BPA concentrations on a terrestrial model organism, the earthworm Eisenia fetida, was assessed. Animals were cutaneously exposed to 100 nM and 10 μM BPA up to 10 days (10-d). Next, a battery of biomarkers was used for ecotoxicological evaluation on a cellular, tissue and behavioural level. HPLC analysis showed that after a 10-d exposure, BPA accumulation reached a maximum of 2.50 μg BPA per g of wet tissue weight. On the cellular level, up to 3-d BPA exposure caused increased lipid oxidation indicating oxidative stress. Histopathological assessment of cell wall and ovaries after 7- and 10-d BPA exposure showed multiple abnormalities, i.e. hyperplasia of epidermis, increased body wall thickness and ovarian atrophy. Detection of these changes was facilitated by a newly proposed semi- quantitative scoring system. Finally, behavioural changes were detected after only 3 days of exposure to 100 nM BPA. Altogether, the presented multilevel toxicity evaluation indicates high sensitivity of earthworms to low BPA doses.

Tolerance towards environmental stress has been frequently considered as one of the key determinants of invasion success. However, empirical evidence supporting the assumption that invasive species can better endure unfavorable conditions compared with native species is limited and has yielded opposing results. In this study, we examined the tolerance to different stress conditions (thermal stress and trace metal zinc pollution stress) in two phylogenetically related and functionally similar freshwater bivalve species, the native Anodonta anatina and the invasive Sinanodonta woodiana. We assessed potential differences in response to stress conditions using several cellular response assays: efficiency of the multixenobiotic resistance mechanism, respiration estimate (INT reduction capacity), and enzymatic biomarkers. Our results demonstrated that the invasive species overall coped much better with unfavorable conditions. The higher tolerance of S. woodiana was evident from (i) significantly decreased Rhodamine B accumulation indicating more efficient multixenobiotic resistance mechanism; (ii) significantly higher INT reduction capacity and (iii) less pronounced alterations in the activity of stress-related enzymes (glutathione-S-transferase, catalase) and of a neurotoxicity biomarker (cholinesterase) in the majority of treatment conditions in both stress trials. Higher tolerance to thermal extremes may provide physiological benefit for further invasion success of S. woodiana in European freshwaters, especially in the context of climate change.

The main goals of this study were (1) to standardize a simple and reliable embryotoxicity test for environmental contaminants and (2) to evaluate the presence and possible protective role of the multidrug resistance-associated (MRP) protein-mediated multixenobiotic defense in two Mediterranean sea urchin species, Paracentrotus lividus and Arbacia lixula. Toxic end-point used was the success of the first cell division in sea urchin embryos. Embryotoxicities of three environmentally relevant contaminants: mercuric chloride (HgCl2, 0.05–6 μM), trybutiltin (TBT, 2.5–500 nM), and oxybenzone (OXI, 0.1–100 μM); as well as seawater samples collected from the polluted and unpolluted locations, were determined and compared. A. lixula embryos were more sensitive to all three toxic compounds, and both P. lividus and A. lixula embryos were highly sensitive to TBT at nanomolar concentrations (EC50 49 ± 5 and 36.8 ± 3 nM, respectively). Inhibition of MRP protein by specific inhibitor MK571 caused significant increase in embryotoxic potency of HgCl2 (EC50 0.697 ± 0.03 and 0.245 ± 0.04 μM, respectively), TBT (EC50 24 ± 3 and 7.4 ± 1 nM, respectively) and polluted seawater sample, but not of OXI or unpolluted, natural seawater. Therefore, our results demonstrated for the first time the protective relevance of MRP proteins in early development of these two Mediterranean sea urchin species. Finally, the embryotoxicity protocol described in this study represents a simple and rapid bioassay for determination of environmentally relevant seawater contamination.

In this study we examined the role of multidrug efflux transport in differential accumulation and potency of inorganic (HgCl2) and organic (CH3HgCl) mercury in sea urchin (Strongylocentrotus purpuratus) embryos. To quantify mercury toxicity we assessed the effect of HgCl2 and CH3HgCl on cell division. Our assay detects the incidence of interference with progression through the first cell cycle, which occurs primarily through the effects of mercury on spindles (microtubules) in metaphase and anaphase. We found that the EC50 concentrations for interference with mitosis are 610 nM for HgCl2 and 124 nM for CH3HgCl. Pharmacological inhibition of MRP/ABCC-type multidrug efflux activity significantly increases the anti-mitotic potency of inorganic mercury, but has virtually no effect on the potency of organic mercury - partial inhibition of the MRP mediated efflux activity with 5 µM MK571, a model inhibitor of MRP efflux transporters, increased the toxicity of HgCl2 2.8-fold. Similarly, inhibition of these transporters increased intracellular accumulation of HgCl2 but has no effect on accumulation on CH3HgCl – 5 µM MK571 caused 2-fold and 20 µM MK571 caused 4-fold increase in intracellular mercury accumulation. Our results point to the MRP mediated efflux transport as an important biological determinant of mercury accumulation and subsequent toxic potency in aquatic organisms.

Mussel (Mytilus galloprovincialis (Lamarck, 1819)) is directly exposed to sea water contamination that elicits significant physiological and cellular re- sponse, although its extent mounted in aquaculture- reared in comparison to wild bivalve populations is scarcely known. Therefore, we have compared contam- ination biomarkers in mussels from reared (Marina farm) and wild, anthropogenically affected site (Vranjic Bay). While predictably, the levels of metals (Cu, Cd, Pb, Zn, Fe, and Hg) in whole bivalve tissues determined by atomic absorption spectrophotometry re- sulted in significantly higher concentrations in wild mussels, accompanied by elevated number of apoptotic cells in gills, the activity of multixenobiotic resistance defense mechanism (MXR), measured as the accumulation rate of model substrate rhodamine B (RB) gave contrasting results. The functional RB assay evidenced a lower MXR efflux activity in the gill tissue of wild mussels, indicating two possible scenarios that willneedfurther focus:(1) persistingseawaterpollution increased cell damage of bivalve gill cells and conse- quently led to leakage of the RB into cytoplasm and dysfunctional MXR efflux in wild mussels; or/and (2) a mixture of different toxic compounds present in Vranjic Bay sea water induced oversaturation of MXR efflux, inducing elevated accumulation of the dye. Consequently, it seems that an efficient physiological functioning of MXR in wild mussels is strongly ham- pered by existence of an unknown quantity of sea water pollutants that may endanger intrinsic organismal de- fense system and lead toward the enhancement of toxicity.

Usage of bisphenol A (BPA) in production of polycarbonate plastics has resulted in global distribution of BPA in the environment. These high concentrations cause numerous negative effects to the aquatic biota, among which the most known is the induction of endocrine disruption. The focus of this research was to determine the effects of two experimentally determined concentrations of BPA (100 nM and 4 µM) on cellular detoxification mechanisms during the embryonic development (2-cell, pluteus) of the rocky sea urchin (Paracentrotus lividus), primarily the potential involvement of multidrug efflux transport in the BPA intercellular efflux. The results of transport assay, measurements of the intracellular BPA and gene expression surveys, for the first time indicate the importance of P- glycoprotein (P-gp/ABCB1) in defense against BPA. Cytotoxic effects of BPA, validated by the immunohistochemistry (IHC) and the transmission electron microscopy (TEM), induced the aberrant karyokinesis, and consequently, the impairment of embryo development through the first cell division and retardation.

Disulfide bonds (SS-bonds) are important posttranslational modifications of proteins. They stabilize a three-dimensional (3D) structure (structural SS-bonds) and have also the catalytic or regulatory functions (redox-active SS-bonds). Although SS-bonds are present in all groups of organisms, no comparative analyses of their frequency in proteins from different domains of life have been made to date. Using the Protein Data Bank (PDB), the number and subcellular locations of SS-bonds in Archaea, Bacteria, and Eukarya have been compared. Approximately three times higher frequency of proteins with SS-bonds in eukaryotic secretory organelles (e.g. endoplasmic reticulum) than in bacterial periplasmic/secretory pathways was calculated. Protein length also affects the SS-bond frequency: the average number of SS-bonds is positively correlated with the length for longer proteins (> 200 amino acids) while for the shorter and less stable proteins (< 200 amino acids) this correlation is negative. Medium size proteins (250 - 350 amino acids) indicated a high number of SS-bonds only in Archaea that could be explained by the need for additional protein stabilization in hyperthermophiles. The results emphasize higher capacity for the SS-bond formation and isomerization in Eukarya when compared to Archaea and Bacteria.

Multixenobiotic resistance (MXR) is an important mechanism of cellular efflux mediated by ATP binding cassette (ABC) transporters that bind and actively remove toxic substrates from the cell. This study was the first to identify ABC transporter P-glycoprotein (P-gp/ABCB1) as a representative of the MXR phenotype in earthworm (Eisenia fetida). The identified partial cDNA sequence of ABCB1 overlapped with ABCB1 homologues of other organisms from 58.5 % to 72.5 %. We also studied the effect of five modulators (verapamil, cyclosporine A, MK571, probenecid, and orthovanadate) on the earthworm’s MXR activity by measuring the accumulation of model substrates rhodamine B and rhodamine 123 in whole body tissue of the adult earthworm. MK571, orthovanadate, and verapamil significantly inhibited MXR activity, and rhodamine 123 turned out to better reflect MXR activity in that species than rhodamine B. Our results show that E. fetida can serve well as a test organism for environmental pollutants that inhibit MXR activity., Mehanizam multiksenobiotičke otpornosti (MXR) prisutan je u mnogim organizmima kao važan stanični detoksikacijski mehanizam. Posredovan je aktivnošću ABC prijenosnika koji vežu i aktivno izbacuju različite toksične tvari iz stanice. U ovom radu dani su podaci o molekularnoj identifikaciji ABC prijenosnika (eksportera) - P-glikoproteina (P-gp/Abcb1), kao jednog od predstavnika MXR fenotipa, u gujavici Eisenia fetida. Određen je djelomični slijed identificiranoga gena Abcb1, njegov predviđeni aminokiselinski slijed uspoređen je s ABCB1 homolozima iz drugih organizama i utvrđena je identičnost od 58,5 do72,5 %. Uz to, istraživali smo učinak pet modulatora (verapamil, ciklosporin, MK571, probenecid, ortovanadat) na aktivnost MXR mehanizma tih gujavica. Kako bismo potvrdili modulirajuće djelovanje istraživanih modelnih inhibitora na MXR mehanizam u E. fetida, mjerili smo akumulaciju modelnih supstrata rodamina B (RB) i rodamina 123 (R123) u tijelu spolno zrelih jedinki gujavica testom izlaganja na filtar papiru. Rezultati su pokazali da svi istraživani modulatori značajno inhibiraju MXR transportnu aktivnost. Naši podaci prvi upućuju na prisutnost P-gp/Abcb1 srodnih gena u gujavici E. fetida. Osim toga, ukazali smo na veliku važnost MXR-a kao specifičnog detoksikacijskog mehanizma koji omogućuje preživljavanje gujavica u onečišćenom okolišu.

Background and Purpose: The signal crayfish Pacifastacus leniusculus (Dana, 1852) is one of the most successful invasive species of crayfish in European freshwaters, an extremely diverse though endangered group of ecosystems. The main goal of this study was to functionally characterize multixenobiotic resistance (MXR) mechanism defense activity in P. leniusculus tissues for the first time. MXR mechanism protects the cell from a wide variety of toxic compounds, and it is mediated by the transport activity of ATP-binding cassette (ABC) proteins. Materials and Methods: MXR transporter activity dye assay was performed by using fluorescent model substrate rhodamine B (RB) in combination with inhibitors of MXR efflux pumps: MK571 and Verapamil, known to inhibit multidrug resistance-associated proteins (MRP) and P-glycoprotein (P-gp), respectively. In this assay, the increase in intracellular fluorescence of the substrate dye, indicates inhibition of MXR efflux protein pumps. The assay was performed in three different tissues (gills, hepatopancreas, tail muscle). Additionally, tissues were exposed to selected heavy metals – mercury (HgCl2) and zinc (ZnCl2), known to occur in open freshwaters as pollutants. Results: Optimal time for RB accumulation in gills and hepatopancreas was determined to be 30 minutes. RB efflux in gills was inhibited by MK571 and in hepatopancreas by Verapamil, suggesting that multidrug resistanceassociated proteins are dominant in gills of P. leniusculus, and P-glycoprotein in hepatopancreas. Finally, inhibitory effect of mercury (HgCl2: 10 and 20 μM) and zinc (ZnCl2: 5–20 μM) on multixenobiotic resistance mechanism activity in gills, and only mercury in hepatopancreas, was detected. Conclusions: The results for the first time demonstrate the presence of multixenobiotic resistance mechanism efflux activity as an important tissue specific defense mechanism in P. leniusculus and provide the basis for future molecular and toxicological studies of this invasive and adaptable species.

Multixenobiotic resistance (MXR) represents an important cellular detoxification mechanism in aquatic organisms as it provides them robustness toward natural and man-made contaminants. Several ABC transporters have major roles in the MXR phenotype – P-gp/ABCB1, MRP1–3/ABCC1–3 and BCRP/ABCG2. In this study, we identified the presence of ABC transporters involved in the MXR mechanism of Arbacia lixula and Paracentrotus lividus. AlABCB1/P-gp, AlABCC3/MRP3, AlABCC9/SUR-like and AlABCG-like transcripts were identified in A. lixula; and PlABCC1/P-gp, PlABCC3/MRP3, PlABCC5/MRP5, and PlABCC9/SUR-like transcripts in P. lividus. For each of the new partial sequences, we performed detailed phylogenetic and identity analysis as a first step toward full characterization and understanding of the ecotoxicological role of these ABC transporters.

U ovom radu, a na temelju zapisa u strucnim casopisima, monografijama i knjigama, prikazujemo razvitak zadarskog zdravstva u razdoblju od završetka Drugog svjetskog rata do danas (1945. – 2008.). Reorganizacija zdravstvene službe iz ratnih na minimalne mirnodopske uvjete rada pocela je stvaranjem stacionara u Bokanjcu 1944. i otvaranjem bolnice 1945. godine. Nova organizacija na pružanju zdravstvene zaštite sastojala se u uspostavljanju zdravstvenih stanica i ambulanti, organizaciji ljekarnicke djelatnosti, održavanju zdravstvenih tecajeva. Otvaranjem Gradsko-kotarske poliklinike 1948. ujedinjena je izvanbolnicka zdravstvena djelatnost, a otvaranjem Doma zdravlja 1952. godine ujedinjena je kurativna i preventivna izvanbolnicka djelatnost. Medicinski centar u Zadru, kao institucionalizirani oblik integralne zdravstvene zaštite, formiran je 1962. godine. Duševna bolnica u Zemuniku, kao Psihijatrijski odjel Zadarske bolnice, otvorena je 1949., a Psihijatrijska bolnica Ugljan, kao azil za duševne bolesnike, 1955. godine. Djelatnost opskrbe lijekovima provodi Ljekarna Zadar od 1962. godine Zavod za zaštitu zdravlja, kao ustanova za potpunu i cjelovitu preventivnu i socijalno-medicinsku zdravstvenu zaštitu, formiran je 1984. godine. Medicinski centar Zadar razdvojen je 1993. godine na Opcu bolnicu i Dom zdravlja. Strucna knjižnica Opce bolnice Zadar, strucni casopis Medica Jadertina, Znanstvena jedinica i Hrvatski lijecnicki zbor – podružnica Zadar omogucili su uvjete za strucni i znanstveni rad zdravstvenih djelatnika. Tijekom Domovinskog rata (1991. – 1995.) zdravstvenu skrb ranjenicima i bolesnicima pružali su djelatnici u Opcoj bolnici, gradskim i terenskim ambulantama i zdravstveni djelatnici angažirani u pokretnim medicinskim ekipama ukljucenim u borbene postrojbe Zbora narodne garde, Hrvatske vojske i Ministarstva unutarnjih poslova. U razdoblju od završetka Drugog svjetskog rata do danas zadarsko zdravstvo je u stalnom usponu u stvaranju strucnih i znanstvenih kadrova i tehnickoj opremljenosti., In this work, which is based on records from professional magazines, monographs and books, we show the development of Zadar health care in the period from the end of the Second World War until the present day (from 1945 to 2008). The reorganization of health care service from war conditions to minimal working conditions in peacetime started with the formation of an in-patient clinic in Bokanjac in 1944 and the opening of a hospital in 1945. The new organization of health care consisted of setting up in-patient clinics and surgeries, organization of pharmaceutical activities, and holding health care courses. The opening of a city-district polyclinic in 1948 united the out-patient health care activity, and the opening of a Health Center in 1952 united the out-patient curative and preventive activities. Medicinski centar (Medical Center) in Zadar as an institutionalized form of integral health care was established in 1962. The Psychiatric Hospital in Zemunik, as Department of Psychiatry of Zadar Hospital was opened in 1949, while Ugljan Hospital of Psychiatry, as an asylum for mental patients, was opened in 1955. The drug supply activity has been carried out by Ljekarna Zadar since1962. The Health Care Institute, as an establishment for the complete and entire prevention and social and medical health care, was established in 1983.. Medicinski centar Zadar was divided in 1998 into General Hospital and Health Center. Zadar General Hospital Scientific Library, the professional magazine Medica Jadertina, Scientific Unit and Croatian Medical Association Branch Office Zadar have rendered possible the conditions for professional and scientific work of health care workers. During the Croatian War of Independence (1991-1995) health care was given to the injured and sick by workers in General Hospital, city and field surgeries and health care workers engaged in moveable medical teams as part of the fighting units of the Croatian National Guard, Croatian Army and Ministry of Internal Affairs. In the period from the end of the Second World War until today, Zadar health care is continually on the rise in creating expert and scientific personnel and technical equipment.

Green bridges are used to decrease highly negative impact of roads/highways on wildlife populations and their effectiveness is evaluated by various monitoring methods. Based on the 3-year monitoring of four Croatian green bridges, we compared the effectiveness of three indirect monitoring methods: track-pads, camera traps and active infrared (IR) trail monitoring system. The ability of the methods to detect different species and to give good estimation of number of animal crossings was analyzed. The accuracy of species detection by track-pad method was influenced by granulometric composition of track-pad material, with the best results obtained with higher percentage of silt and clay. We compared the species composition determined by track-pad and camera trap methods and found that monitoring by tracks underestimated the ratio of small canids, while camera traps underestimated the ratio of roe deer. Regarding total number of recorder events, active IR detectors recorded from 11 to 19 times more events then camera traps and app. 80% of them were not caused by animal crossings. Camera trap method underestimated the real number of total events. Therefore, an algorithm for filtration of the IR dataset was developed for approximation of the real number of crossings. Presented results are valuable for future monitoring of wildlife crossings in Croatia and elsewhere, since advantages and disadvantages of used monitoring methods are shown. In conclusion, different methods should be chosen/combined depending on the aims of the particular monitoring study.