Mussels (Mytilus galloprovincialis) collected at three locations in Boka Kotorska Bay, on the Montenegrin Adriatic coast, were analyzed for the first time by optical and Raman microscopy to detect microplastics (MPs) and other emerging contaminants in their soft tissues. Concentrations of six trace metals (Cu, Zn, Mn, Fe, Cd, and Hg) were also measured in the same samples by atomic absorption spectroscopy. Mussels from a location near the urban area of Kotor were found to exhibit the highest content of MPs and other pollutants originating from anthropogenic sources, while farmed mussels showed higher carotenoid as well as nylon content. The hypothesis of MPs acting as a possible secondary route of trace metals ingress in mussels, a thus far scarcely studied topic, was evaluated based on a comparative analysis of the obtained results. In this context, it was noticed that nylon filaments originating from mussel farming equipment might contribute to higher trace metal content. The results showed that the simultaneous analysis of different contaminants in mussels can be a significant step forward in marine environment pollution monitoring and the assessment of human health risks associated with the consumption of contaminated seafood.

https://www.mdpi.com/2077-1312/9/5/544/pdf

Study of Microplastics and Inorganic Contaminants in Mussels from the Montenegrin Coast, Adriatic Sea

This work contributes to fill one of the gaps regarding nanoplastic interactions with biological systems by producing polyethylene terephthalate (PET) model nanoplastics, similar to those found in the marine environment, by means of a fast top-down approach based on mechanical fragmentation. Their size distribution and morphology were characterized by laser diffraction and atomic force microscopy (AFM). Their autofluorescence was studied by spectrofluorimetry and fluorescence imaging, being a key property for the evaluation of their interaction with biota. The emission spectra of label-free nanoplastics were comparable with those of PET nanoplastics labeled with Nile red. Finally, the suitability of label-free nanoplastics for biological studies was assessed by in vitro exposure with Mytilus galloprovincialis hemolymphatic cells in a time interval up to 6 h. The nanoplastic internalization into these cells, known to be provided with phagocytic activity, was assessed by fluorescence microscopy. The obtained results underlined that the autofluorescence of the model PET nanoplastics produced in the laboratory was adequate for biological studies having the potential to overcome the disadvantages commonly associated with several fluorescent dyes, such as the tendency to also stain other organic materials different from plastics, to form aggregates due to intermolecular interactions at high concentrations with a consequent decrease in fluorescence intensity, and to dye desorption from nanoparticles. The results of the autofluorescence study provide an innovative approach for plastic risk assessment.

https://www.mdpi.com/2079-4991/12/9/1560/pdf?version=1652330031

Autofluorescence of Model Polyethylene Terephthalate Nanoplastics for Cell Interaction Studies

Raman imaging has advanced recently to be able to directly visualise microplastics and even nanoplastics. However, the generated scanning spectrum matrix, akin to a hyperspectral matrix, is challenging to decode. To this end, herein, logic-based, algebra-based, principal component analysis (PCA)-based, and dual-PCA-based algorithms are compared and combined as a PCA/algebra-based algorithm. Specifically, (i) to increase the signal-noise ratio, multiple images that mapped the multiple characteristic peaks of plastics are merged to cross-check each other. The threshold-based logic algorithm is improved by differentiating the intrinsic peak intensity, using an algebra-based algorithm; (ii) PCA can decode the spectrum matrix to generate PCA spectra and PCA images. The PCA spectra can suggest how to further extract information on plastics to merge the corresponding PCA images to enable the capture of microplastics and nanoplastics, by combining and following up with the algebra-based algorithm, called a PCA/algebra-based algorithm; (iii) dual-PCA analysis is employed to guide and extract the multiple PCA spectra towards image merging, to validate the PCA/algebra-based algorithm. We also show that the score percentages of the eigenvalues of PCA can be used to estimate the size amount of the microplastics and nanoplastics in the scanning area, and how to treat the reversed peaks of the PCA spectrum. Overall, the improvement of the algorithms can lead to more effective decoding of the spectrum matrix.

Raman imaging combined with an improved PCA/algebra-based algorithm to capture microplastics and nanoplastics.

Microplastics in fish and sediments from the Montenegrin coast (Adriatic Sea): Similarities in accumulation.

Plastic pollution in the world has led to an abundance of microplastics (MPs) and has been identified as a potential factor that can lead to serious environmental problems, especially in oceans and seas. Information on the current status of MPs pollution along the Montenegrin coast is insufficiently investigated. This study monitors the abundance, distribution, and sources of MPs, and identifies present polymers in the surface sediment of the Montenegrin coast, as well as comparison with previous research.Ten sampling sites along the Montenegrin coast were selected to collect surface sediment samples. The upper layer of sediment (0-5 cm) was collected by a Petite ponar grab. The samples were dried, and density separation was performed using a NaCl solution. The abundance and morphological characteristics of MPs were determined using an optical microscope (DP-Soft software), while FT-IR analysis was done to identify the polymer type.Microplastics were identified in all sediment samples with an average abundance of 307 ± 133 (SD) MPs/kg in dry sediment. The highest abundance of MPs was found in locations in the vicinity of highly populated areas, near wastewater discharges, and areas with high fishing and tourist activities. The most dominant shape types of MPs in all samples were filaments and fragments. The most common colors of MPs were blue and red, while the dominant MPs sizes were 0.1-0.5 mm and 0.5-1.0 mm. Of the eight identified polymers, PP, PE, and PET were the most common.This study reveals MPs characteristics (abundance, distribution, shape type, colors, size, polymers type) in surface sediment along the Montenegrin coast, as well as the most significant sources of MPs pollution, and provides important data for further research on MPs to identify the effects of MPs pollution on the quality, health, and functionality of the marine environment. 

/pdf/distribution-and-characterization-of-microplastics-in-marine-2etx6ev6.pdf

Distribution and characterization of microplastics in marine sediments from the Montenegrin coast

Considering that microplastics are widespread in the marine environment, in this study we evaluated the presence, identify distribution, abundance, shape type, and color of microplastics in surface sediment along the Montenegrin coast, on the Adriatic Sea. These preliminary results provide the first published record of microplastics found in the surface sediment of this area and highlight the importance of microplastics as a component of marine debris. We documented the presence of microplastics at all sampling locations. The identification of polymer types was performed using Fourier-transform infrared (FTIR) spectroscopy, whereby the presence of three polymer types became evident: polypropylene (54.5%), polyethylene (9.7%), and acrylate copolymer (2.0%). Another 22.2% of particles were unidentified polymers, and the remaining 11.5% were non-synthetic materials. The most common shape type of microplastics was filaments (55.5%), followed by granules (26.3%), fragments (14.9%), and films (3.3%). The dominant colors of microplastics followed the order: blue > yellow > red > clear > black > green > blue-white > white. The average abundance of microplastics in all sampling locations was 609 pieces of microplastic/kg of dry sediment. Compared with other studies, the surface sediment of the Montenegrin coast is moderately to highly polluted with microplastics, depending on the examined location.

/pdf/microplastics-in-surface-sediments-along-the-montenegrin-5auuyjfprt.pdf

Microplastics in Surface Sediments along the Montenegrin Coast, Adriatic Sea: Types, Occurrence, and Distribution

This study presents a review of available data on trace element contents in soft tissues of mussels (Mytilus galloprovincialis) collected from the coastal area of Montenegro, starting with the first research in this area (2005–2007) until the recent years. The aim of this review was to compare the results obtained for mussels during different research periods (autumn 2005–spring 2007, autumn 2007–autumn 2009, autumn 2014–autumn 2015) and to assess the human health risks caused by exposure to trace elements via consumption of these mussels. By two approaches for the human health risk assessment (HHRA), considering oral reference doses by the United States Environmental Protection Agency (US EPA) and provisional tolerable intakes by the Joint FAO/WHO Expert Committee on Food Additives (JECFA), it was shown that the levels of Co, Pb, and Cd were the limiting factors for the consumption of mussels from this coastal area during the whole investigated 10-year period. Besides these elements, during the first period As content was the limiting factor for mussel consumption as well. Also, the results obtained for Li via target hazard quotient (THQ) values in the period 2014–2015 are an indicator of possible mussel contamination by this element, which has not been previously studied in M. galloprovincialis. Since the consumption of seafood is one of the primary sources of human exposure to trace elements, the fact that for Boka Kotorska Bay it was noticed a decrease in average concentrations of most elements over time is particularly important.

Trace Elements in Mussels from Montenegrin Coast: A Risk for Human Health

This study presents a review of available data on heavy metal contents in sediments collected along the Montenegrin coast, starting from 2005 to recent years. The aim was to evaluate the pollution level of Montenegrin coastal sediments by determining the concentrations of heavy metals (Fe, Mn, Zn, Cu, Ni, Pb, Cr, Cd, As, and Hg). The pollution status was evaluated using the contamination factor, pollution load index, and geo-accumulation index, as well as the statistical methods, such as cluster analysis (CA). This study showed that concentrations of individual metals at some locations were extremely high. Mean metal concentrations (mg/kg) in sediment samples during whole investigated period were in the following order: Fe > Mn > Zn > Cr > Ni > Pb > Cu > As > Hg > Cd. The calculated contamination factor and pollution load index values indicated enrichment by either natural processes or anthropogenic influences. According to contamination factor (CF), the contents of Zn, Pb, Ni, Cr, and Hg are responsible for very high contamination. Extreme Igeo values were found for Zn, Cu, Pb, and Cd (strongly or strongly to extremely polluted) for the sediment samples from the location Port of Bar in 2011. Also, the location Ada Bojana was characterized as strongly to extremely polluted with Ni in 2019, and Porto Montenegro was extremely polluted with Hg during 2016 and 2019 with the Igeo values of 6.18 and 6.28, respectively. Locations with the highest PLI values in surface sediments were Porto Montenegro (2018–2019) and Port of Bar (2011, 2014). Cluster analysis grouped the examined metals into two main clusters, indicating possible similar origins of the elements within the same cluster.

Ana Perošević-Bajčeta

Papers

Study of Microplastics and Inorganic Contaminants in Mussels from the Montenegrin Coast, Adriatic Sea

Microplastics in Surface Sediments along the Montenegrin Coast, Adriatic Sea: Types, Occurrence, and Distribution

Distribution and characterization of microplastics in marine sediments from the Montenegrin coast

Trace Elements in Mussels from Montenegrin Coast: A Risk for Human Health

Heavy Metals Toxicity in Sediment and the Marine Environment