Human the plastic debris. This material, in the

Human impacts can change marine ecosystem both
directly and indirectly causing, in the first case, overexploitation and loss
of habitat, while, in the latter ones, changes in interactions in the food web
and in the structure of environment (Goulletquer et al. 2014). Coastal areas
are particularly subject to human pressure, since in this zone are located most
of the anthropogenic activities. Human pressure can modify the natural status
of of physical, chemical, and biological components that characterize marine
ecosystem (Reiss et al. 2014). Regular monitoring of sediment quality is an
important pursuant to assessing the possible influence of anthropogenic
pressure on ecosystem quality (Romeo et al. 2015) and give information to
support management in order to reach a Good Environmental Status (GES) in the Marine
Strategy Framework Directive (MSDF) perspective.

Moreover, as reported by
numerous studies (Cozar et al. 2014; Nuelle et al. 2014), the analysis of
marine sediments is important in the evaluation of the emerging pollutants
such as microplastics, that tend to accumulate in the sea-bottom. At present, microplastics
represent a major global concern affecting all world oceans, defined in the
MSFD as D10 into different categories including the plastic debris. This
material, in the environment is subject to a combination of physical,
biological and chemical processes that reduce its structural integrity (Cole et
al. 2011) producing high densities of smaller debris as microplastics (1-5 mm).
As reported in the Guidance on Monitoring of Marin Litter in European Saes
(2013) the monitoring of litter in seafloor cannot consider all coastal areas
because of limited resources, for this reason also opportunistic approach (i.e. data from other research activity
in the harbour) could be used to improve the existing monitoring plans. Recent
studies testify that microplastic can became a threat of biodiversity, becoming
a vector for the introduction of non-native marine species to new habitats on
floating (Barnes 2002; Derraik 2002; Winston 1982). In addition, because of
their size, microplastics are considered bioavailable to organisms throughout
the food web (Thompson et al. 2004). When ingested, plastics release chemicals compound
(nonylphenols, polybrominated diphenyl ethers, phthalates and bisphenol A)
together with adsorbed hydrophobic pollutants (i.e. PCBs, TBT, DBT, MBT). Even if, the use of these pollutants was
bounded, their extensive use in the past and their low water solubility make
them persistent and able to accumulate both in sediments and in biota (Harris
and Wiberg 2002) and are measured at significant
levels in marine ecosystems and marine food webs (D’Alessandro et al. 2016).

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The sea-floor integrity (D6, MSFD) reflects
characteristics of the sea bottom influencing, in particular, the structure and
functioning of communities living on the sea floor (benthic ecosystems). Disturbance
of the bottom may change structure of benthos community damaging mainly sensible
species causing biodiversity loss. Macroinvertebrates, due to their skill to modify
their community patterns in relation to natural and anthropogenic stress(Warwick
1988) , are considered great biondicators of marine ecosystem (Warwick 1993;
Romeo et al. 2015). Actually, a lot of benthic indices based on structure of macrofaunal
communities were created to assess the ecological quality status (EcoQ) to
support data for MSFD, e.g. AMBI (Borja
et al. 2000), M-AMBI (Muxika et al. 2007), BENTIX (Simboura and Zenetos 2002), BOPA
(Dauvin and Ruellet 2007). These indices, based on the subdivision of species
in different ecological groups, return a value of environmental
quality/disturbance status.

The aim of this paper
is to carry out a quality assessment of a high polluted harbour of the Ionian sub-region
and Central Mediterranean Sea through a multidisciplinary approach that
integrate biotic and abiotic parameters, in order to give data to improve the
monitoring plan of the MSFD regarding shallow waters.


Material and methods

Study area

The Augusta site is
located in the MSFD Ionian sub-region of central Mediterranean Sea, in a
harbour area with a high marine traffic activity. This area hosted a variety of
different chemical and petrochemical refining plants, a commercial harbour and
a basis of the Italian Navy and NATO activities (Sprovieri et al. 2007). The harbour
is closed to the South and East by artificial dams. Two main inlets connect
harbour with open sea: the south-east and the east inlet. The basin is
characterised by three different circulation systems: eastern inlet, dominated
by a tidal current with a northward flowing, south eastern inlet, characterized
by flowing parallel to the coast; the northern portion of the basin, instead,
is characterized by shallow seabed and scarcely affected by active currents (Sprovieri et al. 2007;
Romano et al. 2013). Three small rivers
flow in the area, Mulinello in the North and Marcellino and Cantera in the central
part of the bay (Fig. 1). Due to the dangerous contamination of air, seawater,
and marine biota documented in this area, Augusta coastal area has been
included by the Italian Government in the national remediation plan (G.U.R.I.,
L. 426/1998) and evaluated by the World Health Organization as providing a high
environmental risk.


Sampling activities and laboratory analyses

Samples were carried
out from hard and soft bottoms during the summer of 2013 (Table1, Figure1). Soft
bottom samples were collected by means of a Van Veen grab (0.1 m2)
along four transects perpendicular to coastline at three different depths (5,
10 and 20 m). For each sampling site four replicates were carried out, three of
which were used for biological analysis, and one for the environmental
characterization following the methods described in D’Alessandro et al. (2016)
and Romeo et al. (2015). Sediment characterization was carried out according to
Buchanan and Kain (1971) and the percentage of pebble, gravel, sand, silt and
clay was determined according to the ternary Wentworth scale (Wentworth, 1922).

Plastics debris were
classified according to Guidance on Monitoring of Marine Litter in European
Seas (JRC EU, 2013) adapted. Four size classes were identified: microplastics
(1-5 mm), macroplastics (5-10 mm), megaplastics (10-20 mm) and plastics (>
20 mm) (Barnes 2002; Claessens et al. 2011). Microplastics were extracted
according to Alomar et al. 2016 with some modifications. For each sample, 1 Kg
of sediment was dried at 50° C for 48 h and then sieved for 15 min by means stainless
steel sieves with mesh diameter of: 20; 10; 5; 0.5; 0.1 mm. For each fraction,
plastics were extracted by density separation method and then, sediments were
observed under Stereomicroscope (Zeiss Discovery.V8) with optical enhancement
with a maximum magnification of 80x. Accurate precautions have been used to
prevent contamination during all phases of study according to Woodall et al.
2015. Hard bottom samples were carried out by SCUBA diving, scraping a surface
of 400 cm2 from two pillars within refinery. Three samples were
collected at three different depths (0.5; 6.0 and 15.0 m), for a total amount
of 9 samples per pillar.

Chemical analysis
were conducted taking into account contaminants included in the MSFD monitoring
plane and other contaminants of interest taking into account, according
literature, different typologies of anthropogenic impact of the study area (Falandysz
et al. 2006; Fang et al. 2003; Reli? et al. 2016) . Trace elements (Cd, Hg, Pb,
As, Cr) and other elements (Cu, Ni, and Zn) concentrations were determined
through Inductively Coupled Plasma-Mass Spectrometry (ICP-MS; mod. Agilent
Technologies), according to the US-EPA 6020A method. Quantifications of PCBs
(congeners 28, 52, 77, 81, 101, 118, 126, 128, 138, 153, 156, 169, 180) were
conducted following US-EPA 8082A/2007 standard method. PAHs (acenaphthene,
acenaphthylene, anthracene, dibenzo(a,h)anthracene, benzo(b)-fluoranthene,
benzo(a)pyrene, benzo(ghi)perylene, benzo(k)fluoranthene, chrysene,
dibenzo(a,h)anthracene, fluoranthene, indeno(1,2,3,)pyrene, naphthalene,
phenanthrene, pyrene, perylene, acenaphthene) were determined according to
US-EPA Method 8270D. The chemical analyses of butyltins (BT) in the surface
sediments were conducted according to a modified method from Binato et al.
(1998) and Morabito et al. (1995). Tributyltin (TBT), dibutyltin (DBT),
monobutyltin (MBT) and total butyltins (?BT) were determined as described in Romeo
et al. (2015) and D’Alessandro et al. (2016). All results of chemical analysis
were calculated on dry weight (d.w.), trace elements were expressed in µg/Kg,
persistent organic pollutants in mg/Kg, while the butyltins concentrations were
expressed as ng Sn g-1.

In order to characterise the
benthic communities, the main biodiversity indices were calculated: number of
species (S), Shannon’s index (H’), and Pielou’s evenness (J) (Magurran 1991).
Benthic indices (i.e. AMBI, M-AMBI,
Bentix and BOPA) were calculated to evaluate the environmental status on soft
bottom fauna on abundance of each species. AMBI and M-AMBI were calculated by means of AMBI index software
(version 4.0, available at; BOPA index and its relative
environmental quality were calculated using the revisited formula proposed by
Dauvin and Ruellet (2007). 


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