Microbial Interactions between Amylolytic and Non-Amylolytic Lactic Acid Bacteria Strains Isolated during the Fermentation of Pozol.

Pozol is a Mexican beverage prepared from fermented nixtamalized maize dough. To contribute to understanding its complex microbial ecology, the effect of inoculating on MRS-starch pure and mixed cultures of amylolytic Sii-25124 and non-amylolytic W. confusa 17, isolated from pozol, were studied on their interactions and fermentation parameters. These were compared with L. plantarum A6, an amylolytic strain isolated from cassava. Microbial growth, kinetic parameters, amylolytic activity, lactic acid production, and hydrolysis products from starch fermentation were measured. The population dynamics were followed by qPCR. L. plantarum A6 showed higher enzymatic activity, lactic acid, biomass production, and kinetic parameters than pozol LAB in pure cultures. Mixed culture of each pozol LAB with L. plantarum A6 showed a significant decrease in amylolytic activity, lactic acid yield, specific growth rate, and specific rate of amylase production. The interaction between Sii-25124 and W. confusa 17 increased the global maximum specific growth rate (µ), the lactic acid yield from starch (Ylac/s), lactic acid yield from biomass (Ylac/x), and specific rate of lactic acid production (qlac) by 15, 30, 30, and 40%, respectively, compared with the pure culture of Sii-25124. Interactions between the two strains are essential for this fermentation.

Metaproteomic Insights Into the Microbial Community in Pozol.

Pozol is an acidic, refreshing, and non-alcoholic traditional Mayan beverage made with nixtamalized corn dough that is fermented spontaneously. The extensive analysis of the microbiology, biochemistry and metaproteomics of pozol allowed the construction of a comprehensive image of the fermentation system. The main changes in both the substrate and the microbiota occurred in the first 9 h of fermentation. The increase in microorganisms correlated with the drop in pH and with the decrease in the contents of carbohydrates, lipids, and nitrogen, which shows that this stage has the highest metabolic activity. Bacterial proteins were mainly represented by those of lactic acid bacteria, and among them, the proteins from genus Streptococcus was overwhelmingly the most abundant. Yeast proteins were present in all the analyzed samples, while proteins from filamentous fungi increased up to 48 h. The metaproteomic approach allowed us to identify several previously unknown enzyme complexes in the system. Additionally, enzymes for hydrolysis of starch, hemicellulose and cellulose were found, indicating that all these substrates can be used as a carbon source by the microbiota. Finally, enzymes related to the production of essential intermediates involved in the synthesis of organic acids, acetoin, butanediol, fatty acids and amino acids important for the generation of compounds that contribute to the sensorial quality of pozol, were found.

Evaluation of endosulfan degradation capacity by six pure strains isolated from a horticulture soil.

Endosulfan is an organochlorine pesticide included in the Stockholm Convention for Persistent Organic Compounds. The utilization of endosulfan as the sole source of carbon and its mineralization was evaluated using pure strains of Bacillus subtilis, Bacillus pseudomycoides, Peribacillus simplex, Enterobacter cloacae, Achromobacter spanius, and Pseudomonas putida, isolated from soil with historical pesticide use. The consumption of the α isomer of endosulfan by five of the six strains studied was higher than 95%, while B. subtilis degraded only 76% of the initial concentration (14 mg/L). On the other hand, the degradation of the ß isomer was approximately 86% of the initial concentration (6 mg/L) by B. subtilis, P. simplex, and B. pseudomycoides and 95% by P. putida, E. cloacae, and A. spanius. The ability of A. spanius, P. simplex, and B. pseudomycoides to degrade endosulfan has not been previously reported. The production of endosulfan lactone by the Bacillus strains, as well as A. spanius and P. putida, indicated that endosulfan was degraded by the hydrolytic pathway.

Antioxidant and antimicrobial material by grafting of L-arginine onto enzymatic poly(gallic acid).

Microwave-mediated grafting of L-Arg onto naturally derived and stable multiradical poly(gallic acid) (PGAL) in aqueous media has been successfully achieved. This polymeric material has no adverse effect in human cells as there is no hemolytic activity upon MTT and Neutral Red assays. The analytical and computational characterization studies carried out in this study describe a helical molecular structure with random incorporation of L-Arginine pendant groups from PGAL's backbone. The antioxidant properties of the precursor polymer are preserved as proved by the elimination of stable DPPH and hydroxyl radical scavenging, as well as the FRAP and ORAC assays. Regarding the latter, the oxygen radical inhibition is enhanced compared to PGAL, which is attributed to the guanidyl moieties. PGAL-g-L-Arg displays antimicrobial activity against Gram (+) Listeria monocytogenes and Staphylococcus aureus strains with a MIC of 0.8 g/L and a bacteriostatic effect against Gram (-) Escherichia coli. Additionally, scanning electron and confocal fluorescence microscopies as well as crystal violet colorimetric assay demonstrate that the mechanism involved in the bacterial inhibition is related to the formation of porous channels on the membrane, which is discussed according to the helical secondary structure of the polymer and the amino acid guanidyl moieties interacting to bacterial membranes.

Phytochemical Constituents, Antioxidant, Cytotoxic, and Antimicrobial Activities of the Ethanolic Extract of Mexican Brown Propolis.

Propolis is a complex mixture of natural sticky and resinous components produced by honeybees from living plant exudates. Globally, research has been dedicated to studying the biological properties and chemical composition of propolis from various geographical and climatic regions. However, the chemical data and biological properties of Mexican brown propolis are scant. The antioxidant activity of the ethanolic extract of propolis (EEP) sample collected in México and the isolated compounds is described. Cytotoxic activity was evaluated in a central nervous system and cervical cancer cell lines. Cytotoxicity of EEP was evaluated in a C6 cell line and cervical cancer (HeLa, SiHa, and CasKi) measured by the 3-(3,5-dimethylthiazol-2-yl)2,5-diphenyltetrazolium (MTT) assay. The antibacterial activity was tested using the minimum inhibitory concentration (MIC) assay. Twelve known compounds were isolated and identified by nuclear magnetic resonance spectroscopy (NMR). Additionally, forty volatile compounds were identified by means of headspace-solid phase microextraction with gas chromatography and mass spectrometry time of flight analysis (HS-SPME/GC-MS-TOF). The main volatile compounds detected include nonanal (18.82%), α-pinene (12.45%), neryl alcohol (10.13%), and α-pinene (8.04%). EEP showed an anti-proliferative effect on glioma cells better than temozolomide, also decreased proliferation and viability in cervical cancer cells, but its effectiveness was lower compared to cisplatin.

Omics in traditional vegetable fermented foods and beverages.

For a long time, food microbiota has been studied using traditional microbiological techniques. With the arrival of molecular or culture-independent techniques, a strong understanding of microbiota dynamics has been achieved. However, analyzing the functional role of microbial communities is not an easy task. The application of omics sciences to the study of fermented foods would provide the metabolic and functional understanding of the microbial communities and their impact on the fermented product, including the molecules that define its aroma and flavor, as well as its nutritional properties. Until now, most omics studies have focused on commercial fermented products, such as cheese, wine, bread and beer, but traditional fermented foods have been neglected. Therefore, the information that allows to relate the present microbiota in the food and its properties remains limited. In this review, reports on the applications of omics in the study of traditional fermented foods and beverages are reviewed to propose new ways to analyze the fermentation phenomena.

In Vitro Evaluation of the Probiotic and Safety Properties of Bacteriocinogenic and Non-Bacteriocinogenic Lactic Acid Bacteria from the Intestines of Nile Tilapia and Common Carp for Their Use as Probiotics in Aquaculture.

In this study, seven bacteriocinogenic and non-bacteriocinogenic LAB strains previously isolated from the intestines of Nile tilapia and common carp and that showed potent antibacterial activity against host-derived and non-host-derived fish pathogens were assayed for their probiotic and safety properties so as to select promising candidates for in vivo application as probiotic in aquaculture. All the strains were investigated for acid and bile tolerances, transit tolerance in simulated gastrointestinal conditions, for cell surface characteristics including hydrophobicity, co-aggregation and auto-aggregation, and for bile salt hydrolase activity. Moreover, haemolytic, gelatinase and biogenic amine-producing abilities were investigated for safety assessment. The strains were found to be tolerant at low pH (two strains at pH 2.0 and all the strains at pH 3.0). All of them could also survive in the presence of bile salts (0.3% oxgall) and in simulated gastric and intestinal juices conditions. Besides, three of them were found to harbour the gtf gene involved in pH and bile salt survival. The strains also showed remarkable cell surface characteristics, and 57.14% exhibited the ability to deconjugate bile salts. When assayed for their safety properties, the strains prove to be free from haemolytic activity, gelatinase activity and they could neither produce biogenic amines nor harbour the hdc gene. They did not also show antibiotic resistance, thus confirming to be safe for application as probiotics. Among them, Lactobacillus brevis 1BT and Lactobacillus plantarum 1KMT exhibited the best probiotic potentials, making them the most promising candidates.

Lactic Acid Fermentation of Arabinoxylan From Nejayote by Streptococcus infantarius ssp. infantarius 25124 Isolated From Pozol.

Streptococcus infantarius ssp. infantarius 25124 (Sii-25124) is a lactic acid bacterium (LAB) isolated from pozol, a refreshing beverage prepared by suspending fermented nixtamal (a thermal and alkali-treated maize dough) in water. Although Lactobacillus are the predominant strains in fermented doughs, such as sourdoughs, and non-nixtamalized fermented maize foods, the pozol microbiota is markedly different. This may be the result of the nixtamalization process, which could act as a selective force of some strains. Sii-25124 has been reported as the main amylolytic LAB in pozol; starch is the primary carbon source on nixtamal since monosaccharides and disaccharides are lost during nixtamalization; however, non-amylolytic LAB counts are higher than amylolytic LAB in pozol after 24-h fermentation suggesting that another carbon source is being used by the former bacteria. Hemicellulose (arabinoxylan in maize) becomes available via nixtamalization and is subsequently metabolized by LAB. The aim of this work was to determine whether this bacterium is able to use arabinoxylan as the only carbon source in a defined medium containing arabinoxylan extracted from either nejayote (wash water produced during nixtamal preparation), or beechwood xylan. Xylanase activity in the presence of nejayote arabinoxylan (135.8 ± 48.7 IU/mg protein) was higher than that of beechwood (62.5 ± 19.8 IU/mg protein). Other enzymatic activities, such as arabinofuranosidase and acetyl esterase, were also detected, suggesting the adaptation of the bacterium studied to nixtamal dough. It was concluded that Streptococcus infantarius 25124 isolated from pozol was able to use arabinoxylans, which are present in nixtamal dough, so fermentation does not depend exclusively on free sugars and starch.

Quantitative analyses of the bacterial microbiota of rearing environment, tilapia and common carp cultured in earthen ponds and inhibitory activity of its lactic acid bacteria on fish spoilage and pathogenic bacteria.

The present study aimed to evaluate the bacterial load of water, Nile Tilapia and common Carp intestines from earthen ponds, isolate lactic acid bacteria (LAB) and assess their antimicrobial activity against fish spoilage and pathogenic bacteria. Following enumeration and isolation of microorganisms the antimicrobial activity of the LAB isolates was evaluated. Taxonomic identification of selected antagonistic LAB strains was assessed, followed by partial characterisation of their antimicrobial metabolites. Results showed that high counts (>4 log c.f.u ml-1 or 8 log c.f.u g-1) of total aerobic bacteria were recorded in pond waters and fish intestines. The microbiota were also found to be dominated by Salmonella spp., Vibrio spp., Staphylococcus spp. and Escherichia coli. LAB isolates (5.60%) exhibited potent direct and extracellular antimicrobial activity against the host-derived and non host-derived spoilage and pathogenic bacteria. These antagonistic isolates were identified and Lactococcus lactis subsp. lactis was found as the predominant (42.85%) specie. The strains displayed the ability to produce lactic, acetic, butyric, propionic and valeric acids. Bacteriocin-like inhibitory substances with activity against Gram-positive and Gram-negative (Vibrio spp. and Pseudomonas aeruginosa) bacteria were produced by three L. lactis subsp. lactis strains. In this study, the LAB from the microbiota of fish and pond water showed potent antimicrobial activity against fish spoilage or pathogenic bacteria from the same host or ecological niche. The studied Cameroonian aquatic niche is an ideal source of antagonistic LAB that could be appropriate as new fish biopreservatives or disease control agents in aquaculture under tropical conditions in particular or worldwide in general.

Effect of postharvest UV-C treatment on the bacterial diversity of Ataulfo mangoes by PCR-DGGE, survival of E. coli and antimicrobial activity.

Since Mexico is the second largest exporter of mangoes, its safety assurance is essential. Research in microbial ecology and knowledge of complex interactions among microbes must be better understood to achieve maximal control of pathogens. Therefore, we investigated the effect of UV-C treatments on bacterial diversity of the Ataulfo mangoes surface using PCR-DGGE analysis of variable region V3 of 16S rRNA genes, and the survival of E. coli, by plate counting. The UV-C irradiation reduced the microbial load on the surface of mangoes immediately after treatment and the structure of bacterial communities was modified during storage. We identified the key members of the bacterial communities on the surface of fruits, predominating Enterobacter genus. Genera as Lactococcus and Pantoea were only detected on the surface of non-treated (control) mangoes. This could indicate that these genera were affected by the UV-C treatment. On the other hand, the treatment did not have a significant effect on survival of E. coli. However, genera that have been recognized as antagonists against foodborne pathogens were identified in the bands patterns. Also, phenolic compounds were determined by HPLC and antimicrobial activity was assayed according to the agar diffusion method. The main phenolic compounds were chlorogenic, gallic, and caffeic acids. Mango peel methanol extracts (UV-C treated and control mangoes) showed antimicrobial activity against strains previously isolated from mango, detecting significant differences (P < 0.05) among treated and control mangoes after 4 and 12 days of storage. Ps. fluorescens and Ps. stutszeri were the most sensitive.

Growth inhibition of Streptococcus from the oral cavity by α-amyrin esters.

Five terpenoids were tested by the macrodilution broth method to determine their inhibition activity on cariogenic bacterial growth. In general, α-, β-amyrin and α-amyrin phenylacetate proved to be active, reducing the bacterial viability to less than 20%.

Ethnobotanical survey and antibacterial activity of plants used in the Altiplane region of Mexico for the treatment of oral cavity infections.

ETHNOPHARMACOLOGICAL RELEVANCE: The ethnobotanical survey conducted in this study showed 47 plant species used in the Altiplane region of Mexico for the treatment of dental diseases such as toothache, dental caries, periodontal disease and gingivitis. MATERIALS AND METHODS: Information was collected by performing interviews. The following data were recorded: name of the patients or herbalists with their age, sex; date and place of gathering information; pathology of persons interviewed; name of the drug (vernacular name); parts used (leaves, bark, fruits, seeds, aerial parts), mode of preparation and administration, and possible combinations. The inhibitory effects of the aqueous and ethanolic extracts of the medicinal plants detected during the survey the on the growth of Streptococcus mutans and Phrophyromonas gingivalis were determined using microdilution method. The minimum bactericidal concentrations (MBC) were determined from the wells of microplate with no visible bacterial growth. RESULTS: In total, tree places of the Altiplane region of Mexico were visited and five healers and 100 patients were questioned. 47 wild and cultivated species were recorded. The most frequent uses were to treat tooth pain, gum diseases, bad breath and cavities. Infusions were the most frequently prepared formulation. Other applied preparations mentioned with decreasing frequency were decocts, syrups, tinctures, direct application of the plant material without prior preparation and finally macerations. The ethanolic extracts of Haematoxylon brasiletto, Punica granatum, Iostephane heterophyla, Bursera simaruba, Cedrela odorata and Rhus standleyi (12.5-65.0 µg/mL) as well as water extracts of Haematoxylon brasiletto, Punica granatum, Iostephane heterophyla, Amphipterygium adstringens, Argemone mexicana, Cedrela odorata, Eysenhardtia polystachya, Persea americana, Syzygium aromaticum, Cinnamomun zeylanicum, Cnidoscolus multilobus and Rhus standleyi (10.5-78.0 µg/mL) showed the highest inhibitory effect against Streptococcus mutans and Porphyromonas gingivalis. CONCLUSIONS: Many plants are used in the Mexican traditional medicine to treat oral bacterial diseases by the healers or patients. Our study demonstrated that most of the medicinal plants showed an antibacterial effect in vitro, and justified at least in part their use in traditional medicine. These results encourage further investigations to extract and identify the active chemical compounds responsible for the antibacterial effect observed.

Ent-trachyloban-19-oic acid isolated from Iostephane heterophylla as a promising antibacterial agent against Streptococcus mutans biofilms.

From the roots of Iostephane heterophylla, six known compounds, namely, ent-trachyloban-19-oic acid (1), the mixture of ent-kaur-16-en-19-oic acid (2) and ent-beyer-15-en-19-oic acid (3), xanthorrhizol (4), 16α-hydroxy-ent-kaurane (5) and 16α-hydroxy-ent-kaur-11-en-19-oic acid (6) were isolated using a bioassay-guided fractionation method. The known compounds (1-6) were identified by comparison of their spectroscopic data with reported values in the literature. In an attempt to increase the resultant antimicrobial activity of 1 and 4, a series of reactions was performed on ent-trachyloban-19-oic acid (1) and xanthorrhizol (4), to obtain derivatives 1a, 1b, and 4a-4d. All the isolated compounds (1-6) and the derivatives 1a, 1b, and 4a-4d were evaluated for their antimicrobial activity against two oral pathogens, Streptococcus mutans and Porphyromonas gingivalis associated with caries and periodontal disease, respectively. Compounds 1, 1b, 2+3, 4 and 4d inhibited the growth of S. mutans with concentrations ranging from 4.1 µg/mL to 70.5 µg/mL. No significant activity was found on P. gingivalis except for 4 with an MIC of 6.8 µg/mL. The ability of 1, 1b, 2+3, 4 and 4d to inhibit biofilm formation by S. mutans was evaluated. It was found that 1, 1b, 4 and 4d interfered with the establishment of S. mutans biofilms, inhibiting their development at 32.5, 125.0, 14.1 and 24.4 µg/mL, respectively.

[Methods for the study of microbial communities in fermented foods]. / Métodos para el estudio de comunidades microbianas en alimentos fermentados.

The importance of ecological concepts to understand the presence and growth of microorganisms in foods is presently recognized. The production of fermented foods under controlled conditions and its safety assurance depend on the knowledge and control of their microbiota. Traditional fermented foods are obtained by natural fermentations (in which no inoculum is added) and contain complex microbiotas, which are difficult to describe through the use of traditional microbiological methods. The microbial structure of these foods can be studied with different approaches. One of them consists on the typification of isolated microorganisms with methods based on DNA as RFLP, ribotyping, AFLP, ARDRA and RAPD. In order to detect non-culturable or not yet cultured microorganisms, nucleic acids are directly extracted from foods and the microbial diversity is determined from them. Examples of these techniques are the construction of 16S rDNA clone libraries and fingerprinting techniques, such as DGGE and TGGE. Recent advances on the application of these techniques on the study of fermented foods are presented.

Condiciones de salud de familias desplazados por violencia en Urabá, 1997 / Health conditions of displaced families by violence in Uraba, 1997

La investigación de carácter descriptivo profundizó sobre las condiciones de salud de familias desplazadas por violencia, indagó sobre el concepto salud-enfermedad y su influencia en los itinerarios de salud y autocuidado, el acceso a los servicios de salud, la morbilidad referida por la familias y las condiciones de la vivienda, el habitat y el microambiente. Realizando comparaciones entre los resultados según la ubicación de la familia, y observando los cambios antes y después del desplazamiento sobre los mismos temas. La unidad de análisis y observación fue la familia; la muestra poblacional estuvo constituía por 60 familias desplazadas, 30 ubicadas en Urabá y 30 en Medellín. Las familias antes de ser desplazadas utilizan prácticas tradicionales: el cultivo y uso de plantas medicinales para cuidar su salud, problemas de acceso geográfico y social para asistir a los servicios de salud. Posterior al desplazamiento se hallaron diferencias referentes a la asistencia a los servicios de salud y la afiliación a los mismos. Quienes residen en Urabá tienen mejor cobertura en servicios que los ubicados en Medellín. La morbilidad referida antes y después del desplazamiento es estadísticamente significativa, los síntomas referidos al sistema nervioso y al humor cambian drásticamente y evidencian el conflicto de la zona. Medellín ofrece dificultades diferentes pero "alivia" el miedo