Adriana Sturion Lorenzi

Non-ribosomal peptides produced by Brazilian cyanobacterial isolates with antimicrobial activity

Cyanobacterial strains isolated from terrestrial and freshwater habitats in Brazil were evaluated for their antimicrobial and siderophore activities. Metabolites of fifty isolates were extracted from the supernatant culture media and cells using ethyl acetate and methanol, respectively. The extracts of 24 isolates showed antimicrobial activity against several pathogenic bacteria and one yeast. These active extracts were characterized by Q-TOF/MS. The cyanobacterial strains Cylindrospermopsis raciborskii 339-T3, Synechococcus elongatus PCC7942, Microcystis aeruginosa NPCD-1, M. panniformis SCP702 and Fischerella sp. CENA19 provided the most active extracts. The 50 cyanobacterial strains were also screened for the presence of non-ribosomal peptide synthetase (NRPS) and polyketide synthase (PKS) genes and microcystin production. Putative fragment genes coding for NRPS adenylation domains and PKS keto-synthase domains were successfully PCR amplified from 92% and 80% of cyanobacterial strains, respectively. The potential therapeutical compounds siderophores were detected in five cyanobacterial isolates. Microcystin production was detected by ELISA test in 26% of the isolates. Further a protease inhibitor substance was detected by LC-MS/MS in the M. aeruginosa NPLJ-4 extract and the presence of aeruginosin and cyanopeptolin was confirmed by PCR amplification using specific primers, and sequenced. This screening study showed that Brazilian cyanobacterial isolates are a rich source of natural products with potential for pharmacological and biotechnological applications.

Subgeneric diversity of Brasilonema (Cyanobacteria, Scytonemataceae)

The recently described scytonematoid cyanobacterial genus Brasilonema is known mainly from tropical and subtropical rain forests (Mata Atlântica) of southeastern Brazil, where it occurs in aerophytic wooden, stony and iron substrates. This genus was defined according to both molecular and morphological criteria. The type species B. bromeliae was described from the specialized habitat: it grows in phytothelmes, epiphytic on both living and died leaves within the rosettes of large bromeliad plants slightly above or in the zone of the water level. The genus Brasilonema is evidently widely distributed in coastal forests of Sao Paulo State, where it occurs also in remarkable diversity. According to our results, this genus currently comprises seven taxa, which are distinct by different morphology and ecological characteristics.

A NOVEL EPIPHYTIC CYANOBACTERIAL SPECIES FROM THE GENUS BRASILONEMA CAUSING DAMAGE TO EUCALYPTUS LEAVES

A cyanobacterial mat colonizing the leaves of Eucalyptus grandis was determined to be responsible for serious damage affecting the growth and development of whole plants under the clonal hybrid nursery conditions. The dominant cyanobacterial species was isolated in BG‐11 medium lacking a source of combined nitrogen and identified by cell morphology characters and molecular phylogenetic analysis (16S rRNA gene and cpcBA‐IGS sequences). The isolated strain represents a novel species of the genus Brasilonema and is designated Brasilonema octagenarum strain UFV‐E1. Thin sections of E. grandis leaves analyzed by light and electron microscopy showed that the B. octagenarum UFV‐E1 filaments penetrate into the leaf mesophyll. The depth of infection and the mechanism by which the cyanobacterium invades leaf tissue were not determined. A major consequence of colonization by this cyanobacterium is a reduction in photosynthesis in the host since the cyanobacterial mats decrease the amount of light incident on leaf surfaces. Moreover, the cyanobacteria also interfere with stomatal gas exchange, decreasing CO2 assimilation. To our knowledge, this is the first report of an epiphytic cyanobacterial species causing damage to E. grandis leaves.

Cylindrospermopsin induced changes in growth, toxin production and antioxidant response of Acutodesmus acuminatus and Microcystis aeruginosa under differing light and nitrogen conditions

Growing evidence suggests that some bioactive metabolites (e.g. cyanotoxins) produced by cyanobacteria have allelopathic potential, due to their inhibitory or stimulatory effects on competing species. Although a number of studies have shown that the cyanotoxin cylindrospermopsin (CYN) has variable effects on phytoplankton species, the impact of changing physicochemical conditions on its allelopathic potential is yet to be investigated. We investigated the physiological response of Microcystis aeruginosa (Cyanobacteria) and Acutodesmus acuminatus (Chlorophyta) to CYN under varying nitrogen and light conditions. At 24h, higher microcystins content of M. aeruginosa was recorded under limited light in the presence of CYN, while at 120h the lower levels of the toxins were observed in the presence of CYN under optimum light. Total MCs concentration was significantly (p<0.05) lowered by CYN after 120h of exposure under limited and optimum nitrogen conditions. On the other hand, there were no significant (p>0.05) changes in total MCs concentrations after exposure to CYN under high nitrogen conditions. As expected, limited light and limited nitrogen conditions resulted in lower cell density of both species, while CYN only significantly (p<0.05) inhibited the growth of M. aeruginosa. Regardless of the light or nitrogen condition, the presence of CYN increased internal H2O2 content of both species, which resulted in significant (p<0.05) changes in antioxidant enzyme (catalase, peroxidase, superoxide dismutase and glutathione S-transferase) activities. The oxidative stress caused by CYN was higher under limited light and limited nitrogen. These results showed that M. aeruginosa and A. acuminatus have variable response to CYN under changing light and nitrogen conditions, and demonstrate that need to consider changes in physicochemical conditions during ecotoxicological and ecophysiological investigations.

Cyanobacterial community and microcystin production in a recreational reservoir with constant Microcystis blooms

Cyanobacterial blooms are increasing worldwide favored by eutrophic conditions of aquatic ecosystems associated with climatic perturbations. Generally, inland lentic systems are more susceptible to the development of harmful blooms. In the Salto Grande Reservoir (Brazil), Microcystis is the most common bloom-forming genus along with a wide range of co-occurring and less-known cyanobacteria taxa. The cyanobacterial community and microcystin production were studied in Salto Grande Reservoir applying biological, toxicological, and molecular approaches. Thirteen cyanobacterial strains belonging to eight genera were isolated and taxonomically investigated based on morphological traits and phylogenetic analyses of their 16S rRNA gene sequence. The morphotypes identified were, in general, in agreement with their phylogeny. The presence of non-ribosomal peptide synthetase (NRPS) and polyketide synthase (PKS) was investigated using PCR gene amplification, which were detected in 76.9 and 84.4% of the strains, respectively. Positive enzyme-linked immunosorbent assays (ELISA) reactions for microcystins were obtained only from the strain Leptolyngbya sp. CENA129. ELISA and high-performance liquid chromatography (HPLC) analyses of the environmental water samples showed the highest microcystin concentration during the dry and rainy seasons, respectively. This study highlights that microcystin production must be suspected in benthic forms as well as in genera that are morphologically similar but belonging to other evolutionary lineages.

Bioactive Cyanopeptides Produced by Sphaerocavum brasiliense Strains (Cyanobacteria)

The halogenated compounds aeruginosin and cyanopeptolin (protease inhibitors) are well documented in several cyanobacteria. In this work, the presence of genetic coding for aeruginosin and cyanopeptolin in Sphaerocavum genus and their characterization by mass spectrometry and potential toxic effects were investigated. Three strains of Sphaerocavum brasiliense (CCIBt 3094, CCIBt 3096 and CCIBt 3316) isolated from different reservoirs in Sao Paulo State, and one (CCIBt 3179) from Rio Grande do Norte State were selected for this study. Fragments of aerA-aerB and mcnC-mcnE gene regions (aeruginosin and cyanopeptolin, respectively) were amplified by polymerase chain reaction (PCR) and sequenced. Compounds were characterized by mass spectrometry. All the studied strains presented aeruginosin and cyanopeptolin synthetase genes. Mass spectrometry showed the presence of several aeruginosin and cyanopeptolin variants. The Allium test showed that the ethanolic extract promoted toxic and cytotoxic actions in acute and chronic treatments. These naturally occurring protease inhibitors can play an important role in the physiopathology of human diseases.