Clostridium perfringens es un bacilo Gram positivo, esporulado, anaerobio, ampliamente distribuido en la naturaleza, que produce cuatro toxinas principales α, β, ε y ι, las cuales permiten su clasificación en cinco toxinotipos (A-E). Algunas cepas producen una enterotoxina (CPE), codificada por el gen cpe, que causa diarrea en seres humanos y en algunos animales. La presencia de los genes de estas toxinas y la sensibilidad a los antibióticos se determinó en 81 cepas de C. perfringens previamente aisladas y que habían sido mantenidas a -80°C; 20 de suelos, 20 de origen animal, 20 de origen humano y 21 de alimentos cocidos no relacionados con brotes alimentarios. De acuerdo con los resultados de PCR, todas las cepas fueron clasificadas como C. perfringens tipo A, debido a que solo se les detectó el gen de la toxina α, mientras que el gen de la enterotoxina (cpe) se detectó en dos cepas (2.5%) aisladas de alimentos, tal como ha sido descrito en otras regiones del mundo. El 44% de las cepas fue resistente a algún antibiótico; clindamicina (41%), cloranfenicol (25%), penicilina (22%) y metronidazol (20%). En general, las cepas provenientes de suelos presentaron los mayores porcentajes de resistencia a casi todos los antibióticos. El 40% de las cepas de suelo presentó multiresistencia (a tres o más grupos de antibióticos), el 30% de las de origen humano, el 14% de las de alimentos y el 5% de las de origen animal. Las altas tasas de resistencia encontradas podrían deberse al amplio uso de antibióticos como promotores de crecimiento de plantas y animales y esas cepas resistentes podrían actuar como reservorio de genes de resistencia que pueden transferirse entre bacterias de diversos ambientes.

Molecular characterization and antimicrobial resistance of Clostridium perfringens isolates of different origins from Costa Rica. Clostridium perfringens, a Gram positive, spore-forming anaerobe, is widely distributed in nature. Based upon their production of four major toxins α, β, ε and ι, C. perfringens is classified into five toxinotypes (A-E). Some strains produce an enterotoxin (CPE), encoded by the cpe gene, which causes diarrhea in humans and some animals. C. perfringens strains that had been previously isolated and been kept at -80°C were analyzed for the presence of toxin genes and for antimicrobial resistance: 20 from soils,20 from animal, 20 from human origin and 21 from food non related to outbreaks. According to PCR results, all strains were classified as C. perfringens type A, since only α toxin gene was detected, while cpe was detected in two strains (2.5%) isolated from food, as it has been described in other world regions. Antibiotic resistance to at least one antibiotic was detected in 44% of the strains, 41% was resistant to clindamycin, 25% to chloramphenicol, 22% to penicillin and 20% to metronidazole. Soils strains showed the highest resistance percentages to almost all antibiotics. Multiresistance (to three or more antibiotic groups) was detected in the strains from soil (40%), human origin (30%), food (14%) and animal origin (5%). The high resistance rates found may be explained by the widespread use of antimicrobials as growth promoters in plants and animals; also these resistant strains may act as reservoir of resistance genes that may be transferred between bacteria in different environments. Rev. Biol. Trop. 59 (4): 1479-1485. Epub 2011 December 01.

Los alimentos listos para su consumo (ALC) son alimentos procesados que pueden consumirse sin ningún tratamiento térmico adicional, lo que ha incrementado su popularidad. El objetivo de este estudio fue determinar la inocuidad y calidad microbiológica de 90 ALC producidos por pequeñas industrias costarricenses, con el fin de evaluar el riesgo para la salud pública. Se analizaron 26 encurtidos, 18 aderezos, 18 ensaladas, 12 dulces en conserva y 16 antipastos. A cada muestra se le determinó el pH y la presencia por cultivo de indicadores de calidad microbiológica y de patógenos (Salmonella, Listeria monocytogenes, Clostridium perfringens, C. botulinum y Bacillus cereus); además, se investigó por PCR la presencia de genes que codifican por las toxinas de C. botulinum y C. perfringens. Un 37% de las muestras tuvo un nivel de acidez que podría permitir la proliferación de patógenos (pH >4.5). En general, los indicadores de vida útil fueron aceptables, siempre y cuando los ALC se mantengan en condiciones de temperatura y humedad adecuadas. Un 64% de las muestras presentó valores de coliformes totales que sugieren mala higiene en su elaboración (NMP/g >1000), que se confirma con el hallazgo de coliformes fecales en el 56% y que las hace inaceptables para el consumo humano. Todos los cultivos para patógenos fueron negativos, excepto cuatro para B. cereus. No se detectaron toxinas de C. botulinum y solo una muestra fue positiva para el PCR de C. perfringens. Este estudio evidencia una importante contaminación fecal en ALC, una situación indeseable y totalmente prevenible si se practican técnicas adecuadas de manejo de alimentos, de higiene y se presta mayor atención a los puntos críticos de control.

Ready-to-eat (RTE) foods are processed foodstuffs which have gained popularity in recent times because they can be ingested without further thermic treatments. In this work, the microbiological quality and safety of 90 samples of RTE foods manufactured by small Costa Rican industries was determined to evaluate whether they represent a Public Health risk. Twenty-six samples of pickled vegetables, 18 dips, 18 salads, and 12 sweet treats were studied. Each sample was analyzed with regard to its pH, the presence of culturable microbiological quality indicators and recognized foodborne pathogens (Salmonella, Listeria monocytogenes, Clostridium perfringens, C. botulinum, and Bacillus cereus) Selected genes encoding toxins of C. botulinum and C. perfringens were screened by PCR. Thirty-seven percent of the samples had a level of acidity that could allow the growth and proliferation of bacterial pathogens (pH >4.5). The shelf-life indicators were acceptable but only if the RTE foods are kept at adequate conditions of temperature and humidity. Sixty-four percent of the RTE foods had total coliforms values that evidence inadequate hygiene practices during its elaboration (MPN/g >1000). This result was confirmed by the finding of fecal coliforms in 56 % of the samples, which, by the way, are inacceptable for human consumption. All cultures for pathogens were negative, except for 4 samples that contained B. cereus. Toxins of C. botulinum were not detected and one single sample was positive for the PCR for C. perfringens. The elevated degree of fecal contamination detected in the RTE could be prevented by means of good manufacturing practices, better hygiene measures and a deeper attention to critical control points.

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.

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.