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The Study of Bacterioplankton Dynamics in the Berlengas Archipelago (West Coast of Portugal) by Applying the HJ‐Biplot Method

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The Study of Bacterioplankton Dynamics in the Berlengas Archipelago (West Coast of Portugal) by Applying the HJ‐Biplot Method The study of bacterioplankton dynamics in the Berlengas Archipelago (West coast of Portugal) by applying the HJ‐biplot method SUSANA MENDES 1,2, M.J. FERNÁNDEZ‐GÓMEZ 2, M.P. GALINDO‐VILLARDÓN 2, F. MORGADO 3, P. MARANHÃO 1,4, U. AZEITEIRO 4,5 & P. BACELAR‐NICOLAU 4,5 Mendes, S., M.J. Fernández-Gómez, M.P. Galindo-Villardón, F. Morgado, P. Maranhão, U. Azeiteiro & P. Bacelar-Nicolau 2009. The study of bacterioplankton dynamics in the Berlengas Archipelago (West coast of Portugal) by applying the HJ-biplot method. Arquipélago. Life and Marine Sciences 26: 25-35. The relationship between bacterioplankton and environmental forcing in the Berlengas Archipelago (Western Coast of Portugal) were studied between February 2006 and February 2007 in two sampling stations: Berlenga and Canal, using an HJ-biplot. The HJ-biplot showed a simultaneous display of the three main metabolic groups of bacteria involved in carbon cycling (aerobic heterotrophic bacteria, sulphate-reducing bacteria and nitrate-reducing bacteria) and environmental parameters, in low dimensions. Our results indicated that bacterial dynamics are mainly affected by temporal gradients (seasonal gradients with a clear winter versus summer opposition), and less by the spatial structure (Berlenga and Canal). The yearly variation in the abundance of aerobic heterotrophic bacteria were positively correlated with those in chlorophyll a concentration, whereas ammonium concentration and temperature decreased with increasing phosphates and nitrites concentration. The relationship between aerobic heterotrophic bacteria, chlorophyll a and ammonium reveals that phytoplankton is an important source of organic substrates for bacteria. Key words: Berlengas Natural Reserve, aerobic heterotrophic bacteria, sulphate-reducing bacteria, nitrate-reducing bacteria Susana Mendes (e-mail: [email protected]); 1GIRM-Research Group on Marine Resources, Polytechnic Institute of Leiria, School of Tourism and Maritime Technology, Campus 4, PT-2520-641 Peniche, Portugal; 2University of Salamanca, Department of Statistics, ES-37007 Salamanca, Spain; 3CESAM-Centre for Environmental and Marine Studies and Department of Biology, University of Aveiro, PT-3810-193 Aveiro, Portugal; 4IMAR - Institute of Marine Research, University of Coimbra, Department of Zoology, PT-3004-517 Coimbra, Portugal; 5Open University, Department of Sciences and Technology, PT-1269-001 Lisboa, Portugal. The Berlengas Natural Reserve (BNR) is an heritage and to ensure sustainable development of archipelago formed by 3 groups of islands human activities in the area. The Reserve has a (Berlenga, Estelas and Farilhões) located 5.7 total area of 9560 hectares, 9456 of which are miles from Cabo Carvoeiro (Peniche). It was marine. The fauna is characteristic, very rich, created in 1981, aiming to preserve a rich natural diverse and peculiar (Rodrigues et al. 2008). 25 Despite its known biodiversity, the few existing and chemical parameters) that control the annual marine scientific studies were carried out only dynamics of the bacterioplankton in the Berlengas after the establishment of the BNR. Azeiteiro et archipelago area, by an HJ-biplot (Galindo 1986; al. (1997) and Pardal & Azeiteiro (2001) studied Galindo & Cuadras 1986). The HJ-biplot analysis the macrozooplankton distribution and several is an extension of the classical biplots introduced studies addressed the macrobenthos communities by Gabriel in 1971 (Gabriel 1971, 1972; Gabriel (Almeida 1996; Bengala et al. 1997a, 1997b, & Odoroff 1990). The usefulness of HJ-biplot has 1997c; Metelo 1999; Neto 1999; Neto et al. been showed in many fields; for example in 1999). Biology (Martínez-Ruiz et al. 2001; Martínez- Ruiz & Fernández-Santos 2005; Cabrera et al. 2006; Martínez-Ruiz et al. 2007); in Geology (García-Talegón et al. 1999) and Medicine (Pedraz et al. 1985), etc. MATERIALS AND METHODS STUDY SITE AND FIELD PROGRAM The BNR is an archipelago formed by 3 groups of islands (Berlenga, Estelas and Farilhões) located 5.7 miles from Peniche (Portugal). The sea floor consists primarily of irregular hard bottom substrate (i.e. rocks covered with sessile biota, including a variety of algae, sponges, hydrozoans, anemones, crustaceans, sea urchins and Fig. 1. Berlengas Archipelago with its 3 groups tunicates) . of islands and the location of the sampling stations (●). The two sampling sites used were Berlenga (inside BNR, 39º24.70’ N, 9º31.22’ W) and Canal Microbes are a key component of the structure and (39º24.56’ N, 9º26.70’ W) (Fig. 1). Water function of aquatic ecosystems, through their samples were collected monthly from subsurface involvement in the various nutrient biogeochemical water (0.2 to 0.4 m depth) between February 2006 cycles namely in coastal areas (Wetz et al. 2008), and and February. in the NW Iberian margin upwelling system (Barbosa et al. 2001). Until now, no studies have been carried out to DETERMINATION OF PHYSICAL AND CHEMICAL assess this component of the ecosystem in the PARAMETERS BNR. Because of bacterial major role in Samples were analysed in situ for salinity and planktonic microbial food webs (they comprise an temperature (WTW MultiLine P4 portable meter). important share of plankton biomass) and their Water samples for chemical analyses and activities impact on ecosystem metabolism and chlorophyll a quantification were collected and function (Gasol & Duarte 2000) this preliminary immediately stored in the dark at low temperature o study in the BNR has a clear ecological (4 C), until further processing was possible. In relevance. the laboratory these water samples were filtered The present study aimed to investigate the through GF/C filters (1.2 μm pore diameter), for annual dynamics (February 2006 – February quantification of photosynthetic pigments. 2007) of three main metabolic groups of Filtrates were used for determination of nutrient heterotrophic bacteria: aerobic heterotrophic contents. Samples were also analysed in the bacteria (AHB), sulphate-reducing bacteria (SRB) laboratory (in triplicate) for their content in silica, and nitrate-reducing bacteria (NRB), and to nitrate, nitrite, ammonium, phosphate and identify the key factors (among various physical chlorophyll a (Strickland & Parsons 1972). 26 SAMPLING PROCEDURE AND ENUMERATION OF correspondence analysis, factor analysis and VIABLE BACTERIA classical biplots: 1) the distances among row Water samples (1 L) for the enumeration of viable markers are interpreted as an inverse function of bacteria were collected in parallel to those similarities, in such a way that closer markers obtained for physical and chemical analyses. (individuals) are more similar. This property Samples were collected in sterile glass flasks, allows for the identification of clusters of filled to capacity, sealed with gas-tight rubber individuals with similar profiles; 2) the lengths of stoppers and immediately placed on ice until the column markers (vectors) approximate the processing in the laboratory (3 h later). Numbers standard deviation of the variables; 3) the cosines of viable AHB, SRB and NRB were estimated by of the angles among the column vectors MPN method (Bacelar-Nicolau et al. 2003). Eight approximate the correlations among variables in replicate ten-fold dilution series of water samples such a way that small acute angles are associated were prepared in appropriate selective liquid with variables with high positive correlations; media (below) in multiwell plates (Bacelar- obtuse angles near to the straight angle are Nicolau et al. 2003). These were incubated at associated with variables with high negative room temperature, in the dark, for 2 weeks correlations and right angles are associated with (aerobes) or 4 weeks (anaerobes). The presence of non-correlated variables. In the same way, the bacteria was scored positive on the basis of cosines of the angles among the variable markers turbidity or precipitate development, and and the axes (Principal Components) approximate confirmed by microscopic observation. Growth of the correlations between them. For standardized SRB and NRB required strict anaerobic data, these approximate the factor loadings in procedures that were used at all times (Strickland factor analysis and 4) the order of the orthogonal & Parsons 1972). YPG, VMN and the saltwater projections of the row markers (points) onto a Widdel and Back media were used, respectively column marker (vector) approximates the order of for AHB, SRB and NRB (Bacelar-Nicolau et al. the row elements (values) in that column. The 2003). In all media the concentration of NaCl was larger the projection of an individual point onto a adjusted to 30 g.L-1, and pH was adjusted to 7. variable vector is, the more this individual deviates from the average of that variable. Several measurements are essential for a STATISTICAL ANALYSIS correct HJ-biplot interpretation (Galindo 1986; Data were organized in a matrix: eight Galindo & Cuadras 1986). For instance, the environmental variables and the abundance of the relative contribution of the factor to the element is three metabolic groups of bacteria studied (AHB, the relative variability of the variable explained SRB and NRB) at the two sites over the 12 by a factor or dimension. Also, for a point (row or months forming a data matrix of size 24 x 11 (the column) on a factorial plan, the quality of data were normalized by columns to homogenize representation can be defined by adding the two the table). relative contributions of these factors to the The HJ-biplot is an
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