Antimicrobial activity of bacteria isolated from tissue of the coral Palythoa caribaeorum (Zoantharia: Sphenopidae) from Paraíba, Brazil coastal reefs / Actividad antimicrobiana de bacterias aisladas del tejido del coral Palythoa caribaeorum (Zoantharia: Sphenopidae) de los arrecifes costeros de Paraíba, Brasil

Abstract Introduction: The coral-associated bacteria with antimicrobial activity may be important to promote the health of their host through various interactions, and may be explored as a source of new bioactive compounds. Objective: To analyze the antimicrobial activity of bacteria associated with the zoanthid Palythoa caribaeorum from the coral reefs of Carapibus, Paraiba state, Brazil. Methods: The phylogenetic analysis of the bacteria was conducted based on partial sequences of the 16S rRNA gene using molecular and bioinformatics tools. The antimicrobial activity of the 49 isolates was tested against four bacterial strains and one yeast strain: Bacillus cereus (CCT0198), Escherichia coli (ATCC 25922), Staphylococcus aureus (ATCC 25923), Pseudomonas aeruginosa and Candida albicans (ATCC 10231). The antibiosis and antibiogram assays were conducted and the Minimal Inhibitory Concentration (MIC) was determined by the microdilution method. Results: The bacterial isolates belonged to Firmicutes phylum (84 % of the isolates) and the Proteobacteria phylum (16 % of the isolates). Among the 49 isolates five genera were found, with the Bacillus genus being the most abundant (82 % of the isolates), followed by Vibrio (10 %), Pseudomonas (4 %), Staphylococcus (2 %) and Alteromonas (2 %). Antibiosis test revealed that 16 isolates (33 %) showed antimicrobial activity against one or more of five tested reference strains. The highest number of antagonistic bacteria were found in the Bacillus genus (12 isolates), followed by Vibrio (three isolates) and Pseudomonas (one isolate) genera. The B. subtilis NC8 was the only isolate that inhibited all tested strains in the antibiosis assay. However, antibiogram test with post-culture cell-free supernatant of NC8 isolate showed the inhibition of only B. cereus, S. aureus and C. albicans, and the lyophilized and dialyzed material of this isolate inhibited only B. cereus. The lyophilized material showed bacteriostatic activity against B. cereus, with a MIC value of 125 μg/μl, and in the cytotoxicity assay, the hemolysis value was of 4.8 %, indicating its low cytotoxicity. Conclusions: The results show the antimicrobial potential of some bacterial isolates associated with the P. caribaeourum tissue, especially those belonged to Bacillus genus.

Resumen Introducción: La actividad antimicrobiana realizada por las bacterias asociadas con los corales, además de promover la salud de su huésped, representa una fuente para obtener nuevos compuestos bioactivos. Objetivo: Analizar la actividad antimicrobiana de las bacterias asociadas con el zoantario Palythoa caribaeorum de los arrecifes de Carapibus, Paraíba, Brasil. Metodología: El análisis filogenético de la bacterias se realizó con base en secuencias parciales del gen RNAr 16S utilizando herramientas moleculares y de bioinformática. La actividad antimicrobiana de las cepas se probó contra cuatro cepas bacterianas y una cepa de levadura: Bacillus cereus (CCT0198), Escherichia coli (ATCC 25922), Staphylococcus aureus (ATCC 25923), Pseudomonas aeruginosa y Candida albicans (ATCC 10231), utilizando ensayos antibiosis y antibiograma, y la concentración inhibitoria mínima (CIM) que se determinó por el método de microdilución. Resultados: Las cepas bacterianas pertenecían a Firmicutes (84 %) y Gammaproteobacteria (16 %). Entre 49 cepas se encontraron cinco géneros de bacterias: Bacillus, Vibrio, Pseudomonas, Staphylococcus y Alteromonas. Un total de 19 cepas exhibieron actividad antimicrobiana, siendo el género Bacillus el responsable del mayor número de bacterias antagonistas, con 12 cepas positivas en el ensayo de antibiosis y cuatro en la prueba de antibiograma. El mayor número de bacterias antagonistas se encontró en Bacillus (12 aislamientos), seguido por Vibrio (tres aislamientos) y Pseudomonas (un aisladmiento). El NC8, clasificado como Bacillus subtilis, inhibió todas las cepas estándar en el ensayo de antibiosis y las cepas de B. cereus, S. aureus y C. albicans en la prueba de antibiograma. El material liofilizado del B. subtilis NC8 mostró acción bacteriostática contra B. cereus, con un valor de CIM de 125 μg/μl. En la prueba de citotoxicidad, el grado de hemólisis fue del 4.8 % para el material liofilizado a las concentraciones probadas, lo que indica su baja citotoxicidad. Conclusión: Los resultados muestran el potencial antimicrobiano de algunos aislamientos bacterianos asociados al P. caribaeourum, especialmente los pertenecientes al género Bacillus.

Isolation and identification of early marine biofilm-forming bacteria on commercial paint surface

Aims@#To investigate early marine biofilm-forming bacterial diversity on immersed antimicrobial-free commercial paint substratum in seawater. @*Methodology and results@#Total ten bacterial strains were successfully isolated and identified by complete 16S rRNA sequencing. The isolates morphological, biochemical properties, biofilm-forming ability, extracellular polymeric substance (EPS) productivity and components were characterised. The morphological and biochemical characterization of the strains showed strains-specific variation. All isolates were strong biofilm producers with four motile strains being both flat-bottom and air-liquid-interface biofilm producers, while other strains were only air-liquid interface biofilm producer. Based on 16S rRNA, three strains were identified as Marinomonas communis, two were Marinomonas sp., while the rest were Alteromonas litorea, Alteromonas sp., Salinimonas lutimaris, Idiomarine baltica and Bacillus niabensis. The amount of EPS that the isolates produced ranged from 1.95 to 2.89 g/L and productivity of EPS was inversely correlated with the cell biomass. Analysis of the extracted EPS using attenuated total reflectance-fourier transform infrared (ATR-FTiR) showed that all isolates EPS contained carbohydrates, nucleic acid, protein, DNA/RNA and lipid. @*Conclusion, significance and impact of study@#Bacterial diversity in early stages of biofilm on the commercial paint surface was dominated by Gram-negative bacteria from Gammaproteobacteria class. Isolates with superior cell growth showed lowest EPS production. This finding was expected to provide knowledge on distribution of different marine bacterial species in the biofilm on paint coated surfaces which may beneficial to formularize a new antibiofilm paint additive.

Fungicidal activity of volatile organic compounds from Paenibacillus bacteria against Colletotrichum gloeosporioides / Actividad fungicida de compuestos orgánicos volátiles de la bacteria Paenibacillus contra Colletotri chum gloeosporio ides / Atividade fungicida de compostos orgânicos voláteis da bactéria Paenibacillus contra Colletotrichum gloeosporioides

Abstract Some bacteria release volatile organic compounds (VOCs) that can influence the growth of other microorganisms including some pathogens. Identifying bacteria with antifungal activity makes it possible to use such bacteria in the development of biocontrol agents. Thus, the present study focuses on screening VOCs released by eight isolates from Paenibacillus genus, collected at Old Providence and Santa Catalina coral reef (Colombian Caribbean Sea), with antifungal activity against phytopathogenic fungi Colletotrichum gloeosporioides 26B. The VOCs from Paenibacillus sp (PNM-50) showed inhibition rates higher than 50% in the mycelial fungi growth accompanied by macroscopic morphological changes and a reduction in conidiation. In order to identify the VOCs responsible for this antifungal bioactivity, Headspace-Solid Phase Microextraction (HS-SPME) from the bacterial culture was conducted, followed by Gas Chromatography Mass Spectrometry (GC-MS). The chromatographic results revealed a high abundance of VOCs released just by culture media. Once, the difference between VOCs emitted by culture media and bacteria was established, it was possible to make a putative identification of 2-furanmethanol, phenylacetonitrile, and 2,4-dimethylpentanol as possible VOCs responsible for the antifungal activity.

Resumen Algunas bacterias liberan compuestos orgánicos volátiles (COVs) que pueden influir en el crecimiento de otros microorganismos incluyendo algunos patógenos. La identificación de bacterias con actividad antifúngica hace posible el uso de tales bacterias en el desarrollo de agentes de biocontrol. Así pues, en este estudio, se realizó un examen dirigido exclusivamente a los COVs emitidos por ocho aislamientos bacterianos del género Paenibacillus, recolectados en el arrecife de coral de Providencia y Santa Catalina (Mar Caribe colombiano), con actividad antifúngica contra el hongo fitopatógeno Colletotrichum gloeosporioides 26B. Los COVs del aislamiento Paenibacillus sp (PNM-50) mostraron tasas de inhibición superiores al 50% en el crecimiento micelial del hongo, acompañado de cambios morfológicos macroscópicos y una reducción en la conidiación. Para identificar los COVs responsables de esta bioactividad antifúngica, se realizó microextracción en fase sólida del espacio de cabeza (HS-SPME) del cultivo de las bacterias y posterior análisis por cromatografía de gases acoplada a espectrometría de masas (GC-MS). Los resultados cromatográficos revelaron gran abundancia de COVs emitidos por los medios de cultivo. Una vez que se estableció la diferencia entre los COVs emitidos por el medio de cultivo y las bacterias, fue posible identificar tentativamente 2-furanmetanol, fenilacetonitrilo y 2,4-dimetilpentanol como COVs posiblemente responsables de la actividad antifúngica.

Resumo Algumas bactérias liberam compostos orgânicos voláteis (COV) que podem influenciar o crescimento de outros microorganismos, incluindo alguns patógenos. A identificação de bactérias com atividade antifúngica, possibilita seu uso no desenvolvimento de agentes de biocontrole. Neste estudo, foi realizada uma triagem focada nos COV liberados por oito isolados bacterianos do gênero Paenibacillus, coletados nos recifes de coral Old Providence e Santa Catalina (mar do Caribe colombiano), com atividade antifúngica contra fungos fitopatogênicos Colletotrichum gloeosporioides 26B. Os COV de Paenibacillus sp (PNM-50) apresentaram taxas de inibição superiores à 50% sobre o crescimento de fungos miceliais, acompanhadas de alterações morfológicas macroscópicas e redução da conidiação. Para identificar os COV responsáveis por essa bioatividade antifúngica, foi conduzida uma Microextração de Fase Sólida Headspace (HS-SPME) da cultura bacteriana e, em seguida, foi analisada por Cromatografia Gasosa associada à Espectrometria de Massa (GCMS). Os resultados cromatográficos revelaram uma alta abundância de COVs liberados apenas pelos meios de cultura. Uma vez estabelecida a diferença entre os COV emitidos pelos meios de cultura e bactérias, foi possível fazer uma identificação parcial de 2-furanmetanol, fenilacetonitrila e 2,4-dimetilpentanol como possíveis COV responsáveis pela atividade antifúngica.

Primer reporte de bacterias y dinoflagelados marinos luminiscentes del Parque Nacional Isla del Coco, Costa Rica

Introducción: La bioluminiscencia es la capacidad de ciertos organismos para transformar la energía química en energía lumínica mediante varios procesos bioquímicos. Objetivo: El aislamiento e identificación por primera vez de bacterias luminiscentes en agua marina superficial y la identificación de dinoflagelados luminiscentes marinos del Parque Nacional Isla del Coco, Costa Rica. Metodología: Se colectaron muestras de agua marina obtenida por buceo a 20 m y a nivel superficial de 13 sitios en la Isla del Coco, Costa Rica. Por otra parte, se analizaron muestras de fitoplancton colectadas desde la superficie hasta los 30 m de profundudad en los alrededores de 8 sitios de la Isla del Coco, y se determinaron varias especies luminiscentes pertenecientes a los géneros Ornithocercus y Ceratocorys. Resultados: Se logró obtener 7 aislados bacterianos luminiscentes, se identificaron y caracterizaron bioquímicamente mediante una plataforma automatizada (Vitek) con altos niveles de confianza, se ubicaron taxonómicamente dentro del género Vibrio,2 especies: V. alginolyticus y V. parahaemolyticus, además, algunos aislados presentaron resistencia al antibiótico ampicilina y 100% capacidad hemolítica. Esta investigación muestra evidencia de la presencia de especies microscópicas marinas en Isla del Coco, Costa Rica, capaces de presentar el fenómeno de la luminiscencia, por lo que profundizar en su estudio sería relevante en cuanto a la importancia de estos microorganismos en la producción de metabolitos secundarios y como indicadores de floraciones algales nocivas, por lo que se hace necesario realizar más investigación científica para determinar su potencial biotecnológico. Conclusiones: De la misma forma, los resultados obtenidos en esta investigación sugieren expandir las localidades de colecta y aislamientos de microorganismos luminiscentes, acompañado de una caracterización bioquímica y molecular, con el fin de explorar la diversidad microbiana asociada a eventos de luminiscencia y determinar los ambientes en el que estas especies se desarrollan.

Introduction: Bioluminescence is the ability of certain organisms to transform chemical energy into light energy through various biochemical processes. Objective: Isolation and identification for the first time of luminescent bacteria of superficial marine water, and the identification of marine luminescent dinoflagellates of Isla del Coco National Park, Costa Rica. Methods: Samples of seawater obtained by diving at 20 m and at a surface level of 13 sites were collected. On the other hand, phytoplankton samples collected from the surface up to 30 m deep were analyzed in the surroundings of 8 sites of Cocos Island, and several luminescent species belonging to the genera Ornithocercus and Ceratocorys were determined. Results: Seven luminescent bacterial isolates were obtained, they were identified and characterized biochemically by means of an automated platform (Vitek) with high levels of confidence, they were taxonomically located within the genus Vibrio, 2 species: V. alginolyticus and V. parahaemolyticus, in addition, some isolates presented resistance to the antibiotic ampicillin and 100% hemolytic capacity. This research shows evidence of the presence of marine microscopic species in Cocos Island, Costa Rica, capable of presenting the phenomenon of luminescence, so that further study would be relevant in terms of the importance of these microorganisms in the production of metabolites secondary and as indicators of harmful algal blooms, so it is necessary to conduct more scientific research to determine their biotechnological potential. Conclusions: In the same way, the results obtained in this investigation suggest expanding the collection and isolation of luminescent microorganisms, accompanied by a biochemical and molecular characterization, in order to explore the microbial diversity associated with luminescence events and determine the environments in which that these species develop.

Development of a new protocol for freeze-drying preservation of Pseudoalteromonas nigrifaciens and its protective effect on other marine bacteria

Background: Freeze-drying is known as one of the best methods to preserve bacterial strains. Protectant is the key factor affecting the survival rate of freeze-dried strains. In addition, salinity, bacterial suspension concentration, drying time, and other factors can also affect the survival rate of strains to varying degrees. At present, there are relatively few studies on freeze-drying preservation of marine bacteria. In the present study, we performed the freeze-drying protectant screening and optimized the preservation conditions for Pseudoalteromonas nigrifaciens, which is widely distributed in marine environment. The protective effects of the screened protectants were verified by 18 other marine bacterial strains. Results: The results indicated that the combination of 5.0% (w/v) lactose, 5.0% (w/v) mannitol, 5.0% (w/v) trehalose, 10.0% (w/v) skim milk powder, 0.5% (w/v) ascorbic acid and 0.5% (w/v) gelatin was the best choice for the preservation of P. nigrifaciens. The suggested salinity and concentration of initial cell suspension were 10 g/L NaCl and 1.0 × 109 CFU/mL, respectively. Furthermore, stationary-phase cells were the best choice for the freeze-drying process. The highest survival rate of P. nigrifaciens reached 52.8% when using 5­10% (w/v) skim milk as rehydration medium. Moreover, the other 18 marine strains belonging to Pseudoalteromonas, Vibrio, Photobacterium, Planomicrobium, Edwardsiella, Enterococcus, Bacillus, and Saccharomyces were freezedried under the abovementioned conditions. Their survival rates were 2.3­95.1%. Conclusion: Collectively, our results supported that the protectant mixture and parameters were beneficial for lyophilization of marine bacteria

Caracterización fenotípica y molecular, e influencia de medios de cultivo, en el crecimiento y emisión de luz de bacterias del litoral de La Habana, Cuba / Phenotypic and molecular characterization, and influence of culture media, on the growth and light emission of bacteria from the coast of Havana, Cuba

Introducción: El complejo enzimático emisor de luz de las bacterias luminiscentes es una poderosa herramienta bioquímica, con una amplia variedad de aplicaciones, incluyendo el control de la calidad ambiental. Objetivos: Identificar taxonómicamente dos bacterias luminiscentes de las aguas de la plataforma cubana, así como seleccionar los medios de cultivo que favorezcan su crecimiento y luminiscencia. Métodos: La identificación taxonómica de las bacterias luminiscentes se llevó a cabo utilizando métodos tradicionales y moleculares. Cuatro medios de cultivo (LM, Boss, Chalk, ZoBell) fueron evaluados en función de la tasa de crecimiento específico (μ) y la luminiscencia utilizando un espectrofotómetro Genesys 10UV y un espectro fluorómetro Shimadzu RF-5301pc, respectivamente. Resultados: La caracterización bioquímica y fisiológica de los aislamientos de CBM-976 y CBM-992 mostró similitudes con las especies de Vibrio harveyi. El análisis del posicionamiento taxonómico confirmó una alta correspondencia con las cepas de V. harveyi aisladas de entornos acuáticos, utilizando secuencias parciales de los genes 16S rRNA, gyrB y pyrH. Se seleccionaron los medios de cultivo LM y ZoBell por tener una alta tasa de crecimiento específico de las cepas CBM-976 y CBM-992; así como por mostrar altos valores de luminiscencia. Los resultados permitirán profundizar en la caracterización fisiológica y son el punto de partida para el desarrollo de métodos de detección de contaminantes. Conclusiones: La combinación de las características fisiológicas y bioquímicas, así como las técnicas de biología molecular contribuyeron a determinar la posición taxonómica de las cepas CBM-976 y CBM-992 aisladas de las aguas marinas cubanas como Vibrio harveyi. Además, se seleccionaron los medios de cultivo LM y ZoBell como los más adecuados para el crecimiento y la emisión de luminiscencia de ambas cepas.

Introduction: The light-emitting enzyme complex of luminescent bacteria is a powerful biochemical tool, with a wide variety of applications including environmental quality monitoring. Objectives: To identify taxonomically two luminescent bacteria from Cuban shelf waters, as well as select the culture media that favor their growth and luminescence. Methods: The taxonomic location of the luminescent bacteria was carried out using traditional and molecular methods. Four culture media (LM, Boss, Chalk, ZoBell) were evaluated as a function of specific growth rate (μ) and luminescence, using a Genesys 10UV spectrophotometer and a Shimadzu RF-5301pc spectrofluorometer, respectively. Results: Biochemical and physiological characterization of CBM-976 and CBM-992 isolates showed similarities with Vibrio harveyi species. Phylogenetic positioning analysis confirmed a high correspondence with V. harveyi strains isolated from aquatic environments, using partial sequences of 16S rRNA, gyrB and pyrH genes. LM and ZoBell culture media were selected for having a high specific growth rate of CBM-976 and CBM-992 strains, as well as for showing high luminescence values. The results will allow deepening the physiological characterization and are the starting point for the development of contaminant detection methods. Conclusions: The rational combination of physiological and biochemical characteristics, as well as the molecular approach, contributed to determine the taxonomic position of CBM-976 and CBM-992 strains isolated from Cuban marine waters as Vibrio harveyi. Furthermore, LM and ZoBell culture media were selected as the most suitable for growth and luminescence emission for both strains.

Phylogenetic Framework and Metabolic Genes Expression Analysis of Bacteria Isolated from Contaminated Marine Environments of Niger Delta

Aims: To explore the phylogenetic framework of bacteria isolated from contaminated marine environments of Niger Delta and the expression of the metabolic genes coding for aromatic hydrocarbon degradation and surfactant production. Study Design: Nine treatments designs were set up in triplicates containing 25 mL of sterile modified mineral basal medium supplemented with nine marine hydrocarbon degraders incubated at 24°C for 5 days. Three of the set ups were supplemented each with 1 mg /L of xylene, anthracene and pyrene. Place and Duration of Study: Department of Environmental Sciences, University of South Africa, Pretoria, South Africa between September, 2015 to December, 2017. Methodology: A laboratory scale study was carried on six composite samples of the sediment and water samples from the three studied areas using enrichment, screening, selection, characterization, and PCR assays to explore the phylogenetic framework and metabolic genes expression of the marine bacteria for aromatic hydrocarbon degradation and surfactant production. Results: The findings revealed that there was significant abundance of THB (P = .05) more than TCHUB and more xylene degraders than anthracene and pyrene degraders in the sediment and water samples respectively. The phylogenetic correlational analysis revealed that all the nine selected best degraders out of 48 isolates from the studied area were evolutionary related belonging to the genera: Providencia, Alcaligenes, Brevundimonas, Myroides, Serratia, and Bacillus; able to significantly (P = .05) utilize the all the aromatic hydrocarbons. The existence of catabolic and surfactant genes namely catechol dioxygenase (C23O), rhamnolipid enzyme (rhlB) and surfactin /lichenysin enzyme (SrfA3 /LicA3) genes were detected in only four (4) out of the nine (9) marine aromatic degrading bacteria with 881 base pairs sizes. Conclusion: Thus, the study revealed that these bacterial strains especially Serratia marcescens XYL7 might possess metabolic genes for in situ aromatic hydrocarbon degradation and surfactant production.

Production, characterization and antibacterial activity of prodigiosin pigment produced by Pseudoalteromonas rubra BF1A IBRL associated with a marine macroalgae Enteromorpha sp

Aims@#Marine bacteria are a great source of natural pigments, which can be used as colouring agent in food, textile, cosmetics and aquaculture industry to overcome the drawbacks poses by the synthetic pigments. The aim of the study is to identify the potential bio pigment producer, determine the antimicrobial susceptibilities, and characterize the pigment produced. @*Methodology and results@#In this study, the surface attached marine bacteria isolated from the surface of seaweed, Enteromorpha sp. has been identified as Pseudoalteromonas rubra BF1A IBRL through the molecular identification step. This species produced intracellular and extracellular red pigment with antibacterial activity. The susceptible bacteria include Bacillus subtilis, Bacillus cereus, Methicillin Resistant Staphylococcus aureus, Staphylococcus aureus and also Acinetobacter anitratus with inhibition zone ranges from 7.33 to 10.33 mm, whereas Minimum inhibitory concentration (MIC) values ranges from 0.055 to 8.88 mg/mL. The UV/vis analysis indicated that the maximal absorbance of ISO and DE pigment extract were at 531 and 534 nm, respectively. Based on the antimicrobial activity, the extracellular extract poses greater antibacterial activity, thus was selected as the potential pigment extract and were further evaluated. The Thin Layer Chromatography (TLC) profile of the DE extract showed one major band under visible light ((Rf = 0.87) and the bioautography analysis of the pigmented band showed positive activity against both Gram-positive and Gram-negative bacteria. The pigment in DE extract was identified as prodigiosin based on the spectroscopic properties, presumptive test and HPLC analysis. @*Conclusion, significance and impact of study@#This study highlights the dual benefits of the P. rubra BF1A IBRL pigment extract, which exhibited both tinctorial and pharmacology benefits, thus it can be act as colouring agent with own preservative value in food, textile, or cosmetics industries.

Assessment of cultivation parameters influencing the growth, pigment production and anti-MRSA activity of Pseudoalteromonas rubra BF1A IBRL isolated from Malaysian marine environment

Aims@#Pigments are coloured substances that exhibit important characteristics to many industries including food, textile, cosmetics, food, pharmaceutical and also aquaculture industry. Naturally derived pigments from marine bacteria do not only exhibit the tinctorial property but are also known to possess broad range of antimicrobial activities. From the industrial point of view, the necessity to obtain suitable culture conditions for maximum yield of cell growth and pigment production is of utmost importance. @*Methodology and results@#The effect of cultural conditions, including light, pH, temperature, agitation speed and size of inoculum on bioactivity of an epiphytic marine bacteria, Pseudoalteromonas rubra BF1A IBRL was studied using shake flask technology. The antimicrobial activity was determined using the Lorian method. As a result, prodigiosin pigment extract obtained from P. rubra BF1A IBRL showed inhibitory activity against the MRSA strain. Pseudoalteromonas rubra BF1A IBRL produced the highest level of prodigiosin and anti-MRSA activity (P<0.05) in Marine broth at initial pH of 7.6 incubated at dark condition at temperature of 26 °C, agitation speed of 120 rpm and 2% (v/v) (1 × 106 CFU/mL) of inoculums size. @*Conclusion, significance and impact of study@#A high correlation between pigmentation and antibacterial activity were observed anticipating that the pigment has its own antibacterial properties. The above findings supported the fact that epiphytic marine bacteria were fruitful source for pigmented bioactive compounds, and the physical parameters had significantly influence of the pigment production.

Assessment of cultivation parameters influencing the growth, pigment production and anti-MRSA activity of Pseudoalteromonas rubra BF1A IBRL isolated from Malaysian marine environment

Aims@#Pigments are coloured substances that exhibit important characteristics to many industries including food, textile, cosmetics, food, pharmaceutical and also aquaculture industry. Naturally derived pigments from marine bacteria do not only exhibit the tinctorial property but are also known to possess broad range of antimicrobial activities. From the industrial point of view, the necessity to obtain suitable culture conditions for maximum yield of cell growth and pigment production is of utmost importance. @*Methodology and results@#The effect of cultural conditions, including light, pH, temperature, agitation speed and size of inoculum on bioactivity of an epiphytic marine bacteria, Pseudoalteromonas rubra BF1A IBRL was studied using shake flask technology. The antimicrobial activity was determined using the Lorian method. As a result, prodigiosin pigment extract obtained from P. rubra BF1A IBRL showed inhibitory activity against the MRSA strain. Pseudoalteromonas rubra BF1A IBRL produced the highest level of prodigiosin and anti-MRSA activity (P<0.05) in Marine broth at initial pH of 7.6 incubated at dark condition at temperature of 26 °C, agitation speed of 120 rpm and 2% (v/v) (1 × 106 CFU/mL) of inoculums size. @*Conclusion, significance and impact of study@#A high correlation between pigmentation and antibacterial activity were observed anticipating that the pigment has its own antibacterial properties. The above findings supported the fact that epiphytic marine bacteria were fruitful source for pigmented bioactive compounds, and the physical parameters had significantly influence of the pigment production.

Methods for the genetic manipulation of marine bacteria

Genetic manipulation of bacteria is a procedure necessary to obtain new strains that express peculiar and defined genetic determinants or to introduce genetic variants responsible for phenotypic modifications. This procedure can be applied to explore the biotechnological potential associated with environmental bacteria and to utilize the functional properties of specific genes when inserted into an appropriate host. In the past years, marine bacteria have received increasing attention because they represent a fascinating reservoir of genetic and functional diversity that can be utilized to fuel the bioeconomy sector. However, there is an urgent need for an in-depth investigation and improvement of the genetic manipulation tools applicable to marine strains because of the paucity of knowledge regarding this. This review aims to describe the genetic manipulation methods hitherto used in marine bacteria, thus highlighting the limiting factors of the different techniques available today to increase manipulation efficiency. In particular, we focus on methods of natural and artificial transformations (especially electroporation) and conjugation because they have been successfully applied to several marine strains. Finally, we emphasize that, to avoid failure, future work should be carried out to establish tailored methodologies for marine bacteria.

Antibacterial activity of marine bacteria isolated from sponge Xestospongia testudinaria from Sorong, Papua

Objective:To explore secondary metabolite of bacteria-associated Xestospongia testudinaria from Tanjung Kasuari,Sorong,Papua.Methods:The antimicrobial activities of extracts against two Gram-positive bacteria (Staphylococcus aureus) and three Gram-negative bacteria (Pseudomonas aeruginosa,Eschericia coli and Salmonella typhi) were determined by disk diffusion dilution method.Results:The test showed that of 15 isolates of symbiont bacteria,6 isolates were successfully isolated and coded,numcly,Xp 4.1,Xp 4.2,Xp 4.3,Xp 4.4,Xp 4.5 and Xp 4.6.Of the six bacterial isolates,isolated Xp 4.2 was found to have more powerful antibacterial activity than any other isolates of symbiont bacteria.Antibacterial activity assay for the n-hexane soluble fractions,ethyl-acetate soluble fractions,and n-buthanol soluble fractions revealed more powerful anti-bacterial activity than any other soluble fractions.Phytochemical screening showed alkaloid and steroid/triterpenoid,while identification for isolate of Xp 4.2 bacterial showed bacteria.Conclusions:Metabolites of bacterial associated with marine sponge Xestospongia testudinaria promise to be developed into antibacterial agents.

Antibacterial activity of marine bacteria isolated from sponge Xestospongia testudinaria from Sorong, Papua

Objective To explore secondary metabolite of bacteria-associated Xestospongia testudinaria from Tanjung Kasuari, Sorong, Papua. Methods The antimicrobial activities of extracts against two Gram-positive bacteria (Staphylococcus aureus) and three Gram-negative bacteria (Pseudomonas aeruginosa, Eschericia coli and Salmonella typhi) were determined by disk diffusion dilution method. Results The test showed that of 15 isolates of symbiont bacteria, 6 isolates were successfully isolated and coded, namely, Xp 4.1, Xp 4.2, Xp 4.3, Xp 4.4, Xp 4.5 and Xp 4.6. Of the six bacterial isolates, isolated Xp 4.2 was found to have more powerful antibacterial activity than any other isolates of symbiont bacteria. Antibacterial activity assay for the n-hexane soluble fractions, ethyl-acetate soluble fractions, and n-buthanol soluble fractions revealed more powerful anti-bacterial activity than any other soluble fractions. Phytochemical screening showed alkaloid and steroid/triterpenoid, while identification for isolate of Xp 4.2 bacterial showed bacteria. Conclusions Metabolites of bacterial associated with marine sponge Xestospongia testudinaria promise to be developed into antibacterial agents.

Inhibition of marine biofouling by aquatic Actinobacteria and coral-associated marine bacteria

Aims: Biofouling is a common biology phenomenon occuring on ship surface. This phenomenon has become serious threat in marine industries because of great economic loss. Tributyltin has been used to prevent biofouling, but it turned to cause the environmental problem. Therefore, the discovery of alternative environment-friendly compound is necessarily needed. Methodology and results: Five Actinobacteria isolates and fourteen marine bacteria isolates were tested against the biofilm formation of eight biofouling bacteria isolates that isolated from boat surface and the attachment of three biofouling diatoms (Amphora, Navicula, Nitzschia). Actinobacteria CW17 supernatant showed the broad spectrum activity against all fouling bacteria, whereas BC 11-5 supernatant was the only marine bacteria that capable to inhibit biofilm formation of V. neocaledonicus. Moreover, three representative diatoms attachment could be inhibited by the bioactive compounds produced by Actinobacteria and marine bacteria. CW01 supernatant showed broad spectrum and high activity against all three representative diatoms which is very promising. Molecular identification based on 16S rDNA gene sequence showed eight fouling bacteria isolates were biofilm-forming bacteria. Conclusions, significance and impact of study: This research showed aquatic Actinobacteria and coral-associated marine bacteria have the potential to prevent biofouling formation. Further studies are needed to purify and characterize these antibiofouling compounds for environmental application.

Biomineralization processes of calcite induced by bacteria isolated from marine sediments

Biomineralization is a known natural phenomenon associated with a wide range of bacterial species. Bacterial-induced calcium carbonate precipitation by marine isolates was investigated in this study. Three genera of ureolytic bacteria, Sporosarcina sp., Bacillus sp. and Brevundimonas sp. were observed to precipitate calcium carbonate minerals. Of these species, Sporosarcina sp. dominated the cultured isolates. B. lentus CP28 generated higher urease activity and facilitated more efficient precipitation of calcium carbonate at 3.24 ± 0.25 × 10−4 mg/cell. X-ray diffraction indicated that the dominant calcium carbonate phase was calcite. Scanning electron microscopy showed that morphologies of the minerals were dominated by cubic, rhombic and polygonal plate-like crystals. The dynamic process of microbial calcium carbonate precipitation revealed that B. lentus CP28 precipitated calcite crystals through the enzymatic hydrolysis of urea, and that when ammonium ion concentrations reached 746 mM and the pH reached 9.6, that favored calcite precipitation at a higher level of 96 mg/L. The results of this research provide evidence that a variety of marine bacteria can induce calcium carbonate precipitation, and may influence the marine carbonate cycle in natural environments.

Antimicrobial activity and Characterization of Marine bacteria.

Marine bacteria were isolated from seawater was collected from different coastal areas of the Tamilnadu Sea. The antimicrobial activities of these bacteria were investigated. Ethyl acetate extracts of marine bacterial fermentation were screened for antimicrobial activities using the method of agar diffusion. The results showed that 25 strains of the isolates have antimicrobial activity. The proportion of active bacteria associated with isolated from seawater. The active marine bacteria were assigned to the genera Alteromonas, Pseudomonas, Bacillus and Marinobacter. The TLC autobiographic overlay assay implied that the antimicrobial metabolites produced by four strains with wide antimicrobial spectrum were different. These marine bacteria were expected to be potential resources of natural antibiotic products. It can be concluded that isolation of Marine bacterial samples can offer a numbers of microbial strains for sources of new biomolecules from Marine sources.

Isolation and characterization of pigmented bacteria showing antimicrobial activity from Malaysian marine environment

Aims: Natural products play a prominent role in the discovery of leads for the development of drugs in the treatment of human diseases. Much of nature remains to be explored, especially marine and microbial environments. Methodology and results: Fifty-five pigmented marine bacteria were isolated from sponges, seawater, mangrove sediment, sea cucumber and mussel from different coastal area of Malaysia. The antimicrobial activities of these bacteria were investigated by disk diffusion method against pathogenic bacteria. Out of 55 isolates, 18 isolates exhibited antimicrobial activity, which based on morphological characterization, 53% of them were Gram positive and 47% were Gram negative. All active isolates were able to tolerate more than 4% NaCl in the nutrient agar medium that indicated they were autochthonous to marine environment and moderate salt tolerant in nature. Molecular identification of isolates by the strong antimicrobial activities indicates that isolates WPRA3 (JX020764) and SM11-3j belong to genus Serratia and isolate SDPM1 (JQ083392) belongs to genus Zooshikella. Conclusion, significance and impact of study: The results of present study revealed that the active isolates are potential producer of antimicrobial secondary metabolites and might be utilized as drug candidate

Screening, isolation and purification of antibacterial agents from marine actinomycetes.

As marine environmental conditions are extremely different from terrestrial ones, it is surmised that marine actino-mycetes might produce novel bioactive compounds. Hence marine sediments, collected from the coastal areas of Gokharna and Muradeshwara of Karnataka state, were screened Seventeen isolates were obtained on starch-casein agar media by soil dilution technique. However, only six isolates namely ACT-A2, ACT-A3, ACT-A4, ACT-A5, ACT-A7 and ACT-A15 showed significant antibacterial activity against both gram-positive and gram-negative bacteria. Morphological, cultural and biochemical characterization indicated that the isolates belong to Streptomyces genus of Actinomycetes. Further studies were carried out with the most active ACT-A2. Optimization of media, temperature and pH by shake flask fermentation indicated starch-casein, 28°C and 7 to be suitable for ACT-A2. The production of antibiotics began after 24 h, reached maximum at 72 h and maintained at the same level up to 120 h. Ethyl acetate was used to extract antibacterial compounds from the culture filtrate. TLC was done on silica gel using ethyl acetate: methanol (6:4) and direct bioautography has shown the presence of two active substance, one with Rf 0.8 has more activity than the other with Rf 0.4. Further purification is done by column chromatography using a mixture of dicholoromethane and ethyl acetate. The findings from this investigation reveal that the strain ACT-A2 in order exhibited superior antimicrobial activities to other sediment isolates of actinomycetes.

Screening, isolation and purification of antibacterial agents from marine actinomycetes.

As marine environmental conditions are extremely different from terrestrial ones, it is surmised that marine actino-mycetes might produce novel bioactive compounds. Hence marine sediments, collected from the coastal areas of Gokharna and Muradeshwara of Karnataka state, were screened Seventeen isolates were obtained on starch-casein agar media by soil dilution technique. However, only six isolates namely ACT-A2, ACT-A3, ACT-A4, ACT-A5, ACT-A7 and ACT-A15 showed significant antibacterial activity against both gram-positive and gram-negative bacteria. Morphological, cultural and biochemical characterization indicated that the isolates belong to Streptomyces genus of Actinomycetes. Further studies were carried out with the most active ACT-A2. Optimization of media, temperature and pH by shake flask fermentation indicated starch-casein, 28°C and 7 to be suitable for ACT-A2. The production of antibiotics began after 24 h, reached maximum at 72 h and maintained at the same level up to 120 h. Ethyl acetate was used to extract antibacterial compounds from the culture filtrate. TLC was done on silica gel using ethyl acetate: methanol (6:4) and direct bioautography has shown the presence of two active substance, one with Rf 0.8 has more activity than the other with Rf 0.4. Further purification is done by column chromatography using a mixture of dicholoromethane and ethyl acetate. The findings from this investigation reveal that the strain ACT-A2 in order exhibited superior antimicrobial activities to other sediment isolates of actinomycetes.

Production and characterization of exopolysaccharides (EPS) from biofilm forming marine bacterium

Marine water was used for the biofilm forming bacterial isolation. A biofilm forming device was made by using PVC pipes with sterile glass slides. Five morphologically different bacteria were isolated from the scrapping of glass slides from the device after a month period of time. The efficiency of EPS production was checked for all the isolates. Using scanning electron microscope (SEM), the morphology of microbial cells and colonies was studied. The maximum EPS was produced by the strain B3, which was analyzed and characterized by thin layer chromatography (TLC) and Fourier transform infrared spectroscopy (FTIR) spectra. B3 EPS displayed a strong absorption band of - OH at 3415.31 cm-1 and COOH at 1631.48 cm-1 showing it to be polysaccharide.