Growth of Paecilomyces variotii in B0 (diesel), B100 (biodiesel) and B7 (blend), degradation and molecular detection / Crescimento de Paecilomyces variotii em B0 (diesel), B100 (biodiesel) e B7 (mistura), degradação e detecção molecular

AbstractThe introduction of biodiesel to diesel may allow the fuel to be more susceptible to microorganism growth, especially during incorrect storage. To evaluate the effect of adding biodiesel in pure diesel on the growth of Paecilomyces variotii, microcosms containing pure diesel (B0), blend diesel/biodiesel (B7) and pure biodiesel (B100) were used. In microcosm with minimal mineral medium and B0, B7 or B100, after 60 days, the biomass (dry weight) formed at interface oil-water in B7 and B100 was significantly higher when compared to that of B0. Infrared analysis showed reduction of the carbonile fraction in B7 and B100 suggesting formation of intermediate compounds in B7. To monitor possible contamination of fuel storage tank by P. variotii samples were collected and analysed by specific-PCR assay for detection of P. variotii spores in the aqueous phase. This method was able to detect a minimum of 103 spores ml–1, corresponding to 0.0144 ng µl–1 of DNA. Specificity was tested against Aspergillus fumigatus and Pseudallescheria boydii.

ResumoA introdução de biodiesel ao diesel pode permitir que o combustível se torne mais suscetível ao crescimento de microorganismos, especialmente durante o armazenamento incorreto. Para analisar o efeito da adição de biodiesel em diesel puro no crescimento de Paecilomyces variotii, avaliou-se seu desenvolvimento em microcosmos contendo diesel puro (B0), mistura diesel/biodiesel (B7) e biodiesel puro (B100). Em microcosmos com meio mineral mínimo e B0, B7 ou B100, após 60 dias, a biomassa (peso seco) formada na interface óleo-agua com B7 e B100 foi significativamente maior quando comparada com a de B0. A análise de infravermelho mostrou redução da fração carbonila em B7 e B100, sugerindo a formação de compostos intermediários em B7. Para monitorar uma possível contaminação de tanque de armazenamento de combustível por P. variotii, amostras foram colhidas e analisadas por um teste de PCR específico para detecção de esporos deste fungo em fase aquosa. Este método foi capaz de detectar um mínimo de 103 esporos ml–1, correspondente a 0.0144 ng µl–1 de DNA. Especificidade foi testada contra Aspergillus fumigatus e Pseudallescheria boydii.

Bacterial contamination of automotive fuels in a tropical region: the case of Costa Rica

Microbial contamination of fuel has been the cause of several problems in transportation and storage of these products. Due to the lack of previous studies related to these problems in Costa Rica, bacterial quality was evaluated biannually in automotive fuels stored in the four oil distribution facilities of the Costa Rican Petroleum Refinery (RECOPE). In 12 oil storage tanks, for a total of 96 samples, mesophilic, heterotrophic aerobic/facultative counts (ASTM D6974-04) and identification of bacteria presented in regular gas, premium gas and diesel from the bottom and superior part of the tanks were done; in the samples containing an aqueous phase, sulfate reducing bacteria (SRB) were also quantified by the most probable number technique (MPN), according to the ASTM D4412-84 standard. The higher contamination was shown at the bottom of the tanks (populations up to 10(4) UFC/l), especially if there was accumulated water, in which case populations reached 10(8) UFC/l. The most contaminated fuel was diesel (counts up to 10(4) UFC/l), whereas the less contaminated was premium gas. The less contaminated fuels were from the facilities of La Garita and Barranca, whereas the most contaminated were from Ochomogo. Nevertheless, the quantified populations did not cause significant alteration in quality physicochemical parameters in the samples analyzed. A total of 149 bacterial strains were isolated, 136 (91.3%) Gram positive and 13 (8.7%) Gram negative. The most frequent genera were Staphylococcus (24.0%), Micrococcus (21.9%), Bacillus (18.8%) and Kocuria (11.5%) among Gram positive bacteria and Pseudomonas (7.3%) among Gram negative bacteria. The majority of these genera have been found as fuel contaminants or even as degraders of this kind of products; nevertheless, some species for which their appearance or growth in hydrocarbons have not been described were found with low frequencies. SRB were present in counts up to 10(5) MPN/l in 42.9% of water containing samples (including all from diesel tanks), indicating biocorrosion processes risk in fuel transport and storage systems. From the findings in this study it is recommended to give a frequent maintenance to fuel containers, based on continuous drainage and removal of accumulated water, antimicrobial agent addition and microbial quality monitoring in country’s fuels. Rev. Biol. Trop. 57 (3): 489-504. Epub 2009 September 30.

Se evaluó semestralmente durante dos años la calidad bacteriana de los combustibles almacenados en los cuatro planteles de la Refinadora Costarricense de Petróleo (Costa Rica). Para un total de 96 muestras se realizaron recuentos (norma ASTM D6974-04) e identificación de las bacterias presentes en gasolina regular, gasolina súper y diesel en los niveles superior e inferior de los tanques contenedores; en las muestras con fase acuosa se cuantificaron las bacterias reductoras de sulfato (SRB, norma ASTM D4412-84). La mayor contaminación se observó en el fondo de los tanques, sobretodo si se presentó una capa de agua almacenada. El diesel fue el combustible más contaminado, sin embargo, no se observó alteración importante en los parámetros fisicoquímicos de las muestras evaluadas. Se aislaron 149 cepas, 136 (91.3%) Gram positivas y 13 (8.7%) Gram negativas; los géneros más frecuentes fueron Staphylococcus (24.0%), Micrococcus (21.9%), Bacillus (18.8%), Kocuria (11.5%) y Pseudomonas (7.3%). Con bajas frecuencias se encontraron algunas especies para las que no se ha descrito su aparición o crecimiento en hidrocarburos. Las SRB se presentaron con recuentos de hasta 105 MPN/l en un 42.9% de las muestras con agua residual, principalmente en diesel, lo cual es indicativo de alerta por biocorrosión. A partir de los resultados se recomienda dar un mantenimiento frecuente a los contenedores, la adición de compuestos antimicrobianos y el monitoreo de la calidad microbiana de los combustibles del país.

Natural bioremediation of aquifer material contaminated with gasoline-ethanol mixtures

A biorremediaçäo natural, ou seja, o uso de microorganismos nativos para degradar substâncias periosas sem nenhuma estimulaçäo tecnológica, mostra-se como uma técnica promissora e de baixo custo para se aplicada no manejo de plumas de hidrocarbonetos de petróleo. Esta técnica exige que se faça uma total caracterizaçäo e monitoramento do local contaminado para se verificar se os processos de atenuaçäo natural continuam proporcionando uma adequada proteçäo de riscos. Um grande progresso tem sido feito na Europa e nos Estados Unidos para entender os fatores hidrogelógicos e microbiológicos que influenciam a biorremediaçäo natural de aquíferos contaminados por combustíveis. No entanto, esta experiência deve ser extrapolada com cuidado em locais contaminados no Brasil, onde a gasolina contém 22 por cento de etanol. Estudos preliminares de laboratório mostram que o etanol pode aumentar a solubilizaçäo dos compostos BTX. É necessário que se conheça os efeitos bioquímicos, físicos e ecológicos do etanol para que se desenvolva no Brasil uma base racional para seleçäo, modelagem matemática e monitoramento de sistemas apropriados de biorremediaçäo natural