Co-cultivation of sake yeast and Kocuria isolates from the sake brewing process.

Kocuria isolates collected from the sake brewing process have inhabited the Narimasa Sake Brewery in Toyama, Japan. To investigate the effect of these actinobacterial isolates on the growth and metabolism of sake yeast, co-cultivation of sake yeast and Kocuria isolates was performed in a medium containing tryptone, glucose and yeast extract (TGY), and a solution containing koji (steamed rice covered with Aspergillus oryzae) and glucose. In the TGY medium, the ethanol concentration and the number of living cells of each microorganism were measured. In the koji solution, the concentrations of ethanol and organic acids (citric acid, lactic acid and succinic acid) were measured. The results showed that in TGY media, the growth of each Kocuria isolate in the co-culture of the two Kocuria isolates was similar to that in each monoculture. However, the growth of both Kocuria isolates was inhibited in the co-cultures of sake yeast and Kocuria isolates. On the other hand, the growth and ethanol productivity of sake yeast did not differ between its monoculture and co-cultures with Kocuria isolates. In the koji solution, Kocuria isolates TGY1120_3 and TGY1127_2 affected the concentrations of ethanol and lactic acid, respectively. Thus, Kocuria isolates affected the microbial metabolism, but the effects were not identical between the two isolates. This strongly suggests that bacteria inhabiting a sake brewery may influence the flavor and taste of sake products of the brewery.

Optimization of Fermentation Medium for Indole Acetic Acid Production by Pseudarthrobacter sp. NIBRBAC000502770.

Indole acetic acid (IAA) has been an important compound for plant growth and is widely known to be produced by plant growth-promoting rhizobacteria (PGPR). The isolate producing the maximum amount of IAA from the Korea shooting range soil was identified as Pseudarthrobacter sp. NIBRBAC000502770, using 16S rRNA gene sequencing. IAA production was determined in Luria-Bertani (LB) broth and optimized using different temperatures, agitation rates, L-tryptophan concentrations, carbon and nitrogen sources, and inorganic salts. The strain NIBRBAC000502770 showed better production of IAA at temperature 30 °C (29.47 mg·L-1) and at an agitation rate of 200 rpm (32.65 mg·L-1). Maltose (0.5%) was found to be the best carbon source for the strain (yielding 36.48 mg·L-1 IAA). IAA yield was 19.17 mg·L-1 and 24.73 mg·L-1 at 1% yeast extract and 1% tryptone as nitrogen sources, respectively. qRT-PCR showed the transcript levels of amiE and aldH genes, which had been predicted to encode indole-3-acetamide hydrolase and indole-3-acetaldehyde dehydrogenase, to be significantly upregulated in response to tryptophan. This study has examined that NIBRBAC000502770 has significant effects as a biological agent such as plant growth promotion, and development of optimal medium could significantly reduce the cost of mass production of microorganisms.

Phenotypic heterogeneity as key factor for growth and survival under oligotrophic conditions.

Productivity-poor oligotrophic environments are plentiful on earth. Yet it is not well understood how organisms maintain population sizes under these extreme conditions. Most scenarios consider the adaptation of a single microorganism (isogenic) at the cellular level, which increases their fitness in such an environment. However, in oligotrophic environments, the adaptation of microorganisms at population level - that is, the ability of living cells to differentiate into subtypes with specialized attributes leading to the coexistence of different phenotypes in isogenic populations - remains a little-explored area of microbiology research. In this study, we performed experiments to demonstrate that an isogenic population differentiated to two subpopulations under low energy-flux in chemostats. Fluorescence cytometry and turnover rates revealed that these subpopulations differ in their nucleic acid content and metabolic activity. A mechanistic modelling framework for the dynamic adaptation of microorganisms with the consideration of their ability to switch between different phenotypes was experimentally calibrated and validated. Simulation of hypothetical scenarios suggests that responsive diversification upon a change in energy availability offers a competitive advantage over homogenous adaptation for maintaining viability and metabolic activity with time.

Fermentation conditions optimization, purification, and antioxidant activity of exopolysaccharides obtained from the plant growth-promoting endophytic actinobacterium Glutamicibacter halophytocola KLBMP 5180.

In this study, an endophytic actinobacterium Glutamicibacter halophytocola KLBMP 5180, was investigated for the production and antioxidant activity of exopolysaccharides (EPSs). First, the suitable fermentation time, temperature, inoculation volume, pH value, and the carbon and nitrogen sources for EPSs production were obtained using the one variable at a time method (OVAT). Then, a central composition design was used for fermentation conditions optimization to obtain the maximum EPS yield. The optimal medium and condition were as follows: 100 mL broth in 250 mL Erlenmeyer flasks, including 3.65 g/L maltose, 9.88 g/L malt extract, 3.40 g/L yeast extract, 1.41 g/L MnCl2, pH 7.5, culture temperature 28 °C, and 200 rpm for 7 days, which increased the yield of EPSs to 2.89 g/L. Two purified EPSs, 5180EPS-1 (MW 58.9 kDa) and 5180EPS-2 (10.5 kDa), comprising rhamnose, galacturonic acid, glucose, glucuronic acid, xylose, and arabinose, were obtained for chemical analysis and antioxidant evaluation. The scavenging ability and reducing power of the superoxide anion and hydroxyl radicals demonstrated the moderate in vitro antioxidant activities of the two EPSs, thus indicating their potential to be a new source of natural antioxidants. However, further structure elucidation and functional studies need to be continued.

Euonymus japonicus phyllosphere microbiome is significantly changed by powdery mildew.

Euonymus japonicus Thunb. is a woody and ornamental plant popular in China, Europe and North America. Powdery mildew is one of the most serious diseases that affect E. japonicus growth. In this study, the diseased and apparently healthy leaves were collected from E. japonicus planted in a greenbelt in Beijing, and the effect of powdery mildew on the epiphytic microbial community was investigated by using Illumina sequencing. The results showed that the healthy leaves (HL) harbored greater bacterial and fungal diversity than diseased leaves (DL). Furthermore, both bacterial and fungal communities in DL exhibited significantly different structures from those in HL. The relative abundance of several bacterial phyla (Proteobacteria and Firmicutes) and fungal phyla (Ascomycota and Basidiomycota) were altered by powdery mildew. At the genus level, most genera decreased as powdery mildew pathogen Erysiphe increased, while the genera Kocuria and Exiguobacterium markedly increased. Leaf properties, especially protein content was found to significantly affect beta-diversity of the bacterial and fungal community. Network analysis revealed that positive bacterial interactions in DL were stronger than those in HL samples. Insights into the underlying the indigenous microbial phyllosphere populations of E. japonicus response to powdery mildew will help in the development of methods for controlling plant diseases.

Infection, Survival, and Growth of Clavibacter nebraskensis on Crop, Weed, and Prairie Plant Species.

Clavibacter nebraskensis is the causal agent of Goss's leaf blight and wilt, an important disease of maize in the United States and Canada. The epidemiology and ecology of this bacterial pathogen are poorly understood. Infested maize residue is often considered to be the primary source of inoculum for maize; however, the potential for many other plant species to be infected and serve as inoculum sources is unknown. The goal of this study was to determine if C. nebraskensis could infect, survive, and grow on common weed, crop, and grass species. Seedling leaves of 18 plant species that grow in maize production areas in the United States were inoculated with this pathogen in a controlled environment and in the field. Lesion development, bacterial streaming, and pathogen population size on leaves were then determined and used as criteria to evaluate host-pathogen interactions. Woolly cupgrass (Eriochloa villosa) and the native prairie grasses big bluestem (Andropogon gerardii) and little bluestem (Schizachyrium scoparium) developed lesions and bacterial streaming, identifying them as hosts and susceptible to infection. To our knowledge, this is the first report of these grass species being hosts of C. nebraskensis. Ten other grass species, including wheat and oats, were identified as potential sustaining hosts that maintained epiphytic or endophytic pathogen populations >106 colony forming units per leaf sample but displayed no evidence of infection. Five broadleaf species tested were nonhosts based on the three criteria. This study suggests that multiple plant species support infection and growth of C. nebraskensis and further elucidates the ecology of this pathogen and the epidemiology of Goss's wilt.

Impacts of indoor surface finishes on bacterial viability.

Microbes in indoor environments are constantly being exposed to antimicrobial surface finishes. Many are rendered non-viable after spending extended periods of time under low-moisture, low-nutrient surface conditions, regardless of whether those surfaces have been amended with antimicrobial chemicals. However, some microorganisms remain viable even after prolonged exposure to these hostile conditions. Work with specific model pathogens makes it difficult to draw general conclusions about how chemical and physical properties of surfaces affect microbes. Here, we explore the survival of a synthetic community of non-model microorganisms isolated from built environments following exposure to three chemically and physically distinct surface finishes. Our findings demonstrated the differences in bacterial survival associated with three chemically and physically distinct materials. Alkaline clay surfaces select for an alkaliphilic bacterium, Kocuria rosea, whereas acidic mold-resistant paint favors Bacillus timonensis, a Gram-negative spore-forming bacterium that also survives on antimicrobial surfaces after 24 hours of exposure. Additionally, antibiotic-resistant Pantoea allii did not exhibit prolonged retention on antimicrobial surfaces. Our controlled microcosm experiment integrates measurement of indoor chemistry and microbiology to elucidate the complex biochemical interactions that influence the indoor microbiome.

Specific growth inhibitors of Ralstonia solanacearum, Xanthomonas oryzae pv. oryzae, X. campestris pv. campestris, and Clavibacter michiganensis subsp. michiganensis.

Plant pathogenic bacteria cause huge yield losses in crops globally. Therefore, finding effective bactericides to these pathogens is an immediate challenge. In this study, we sought compounds that specifically inhibit the growth of Ralstonia solanacearum. As a result, we identified one promising compound, 1-(4-bromophenyl)-6-methoxy-2,3,4,9-tetrahydro-1H-ß-carboline, which inhibited the growth of R. solanacearum (Rs1002) from a pilot library of 376 chemicals provided from RIKEN. We further obtained its structural analogues and assessed their ability to inhibit Rs1002 growth. Then we identified five compounds, named ralhibitins A to E, that specifically inhibit growth of Rs1002 at >5 µg/ml final concentration. The most effective compounds, ralhibitins A, C, and E completely inhibited the growth of Rs1002 at 1.25 µg/ml. In addition, ralhibitins A to E inhibited growth of Xanthomonas oryzae pv. oryzae but not the other bacteria tested at a final concentration of 10 µg/ml. Whereas, ralhibitin E, besides inhibiting R. solanacearum and X. oryzae pv. oryzae, completely inhibited the growth of X. campestris pv. campestris and the Gram-positive bacterium Clavibacter michiganensis subsp. michiganensis at 10 µg/ml. Growth inhibition by these compounds was stable at pH 6-9 and after autoclaving. Because Rs1002 grew in the culture medium in which ralhibitins were incubated with the ralhibitin-insensitive bacteria, the unaffected bacteria may be able to inactivate the inhibitory effect of ralhibitins. These results suggest that ralhibitins might be potential lead compounds for the specific control of phytopathogenic bacteria.

Degradation of phenol via ortho-pathway by Kocuria sp. strain TIBETAN4 isolated from the soils around Qinghai Lake in China.

Based on the feature of high-altitude permafrost topography and the diverse microbial ecological communities of the Qinghai-Tibetan Plateau, soil samples from thirteen different collection points around Qinghai lake were collected to screen for extremophilic strains with the ability to degrade phenol, and one bacterial strain recorded as TIBETAN4 that showed effective biodegradation of phenol was isolated and identified. TIBETAN4 was closely related to Kocuria based on its observed morphological, molecular and biochemical characteristics. TIBETAN4 grew well in the LB medium at pH 7-9 and 0-4% NaCl showing alkalophilicity and halophilism. The isolate could also tolerate up to 12.5 mM phenol and could degrade 5 mM phenol within 3 days. It maintained a high phenol degradation rate at pH 7-9 and 0-3% NaCl in MSM with 5 mM phenol added as the sole carbon source. Moreover, TIBETAN4 could maintain efficient phenol degradation activity in MSM supplemented with both phenol and glucose and complex water environments, including co-culture Penicillium strains or selection of non-sterilized natural lake water as a culture. It was found that TIBETAN4 showed enzymatic activity of phenol hydroxylase and catechol 1,2-dioxygenase after induction by phenol and the corresponding genes of the two enzymes were detected in the genome of the isolate, while catechol 2,3-dioxygenase or its gene was not, which means there could be a degradation pathway of phenol through the ortho-pathway. The Q-PCR results showed that the transcripts of both the phenol hydroxylase gene and catechol 1,2-dioxygenase gene were up-regulated under the stimulation of phenol, demonstrating again that the strain degraded phenol via ortho-degradation pathway.

Tracking Polymicrobial Metabolism in Cystic Fibrosis Airways: Pseudomonas aeruginosa Metabolism and Physiology Are Influenced by Rothia mucilaginosa-Derived Metabolites.

Due to a lack of effective immune clearance, the airways of cystic fibrosis patients are colonized by polymicrobial communities. One of the most widespread and destructive opportunistic pathogens is Pseudomonas aeruginosa; however, P. aeruginosa does not colonize the airways alone. Microbes that are common in the oral cavity, such as Rothia mucilaginosa, are also present in cystic fibrosis patient sputum and have metabolic capacities different from those of P. aeruginosa Here we examine the metabolic interactions of P. aeruginosa and R. mucilaginosa using stable-isotope-assisted metabolomics. Glucose-derived 13C was incorporated into glycolysis metabolites, namely, lactate and acetate, and some amino acids in R. mucilaginosa grown aerobically and anaerobically. The amino acid glutamate was unlabeled in the R. mucilaginosa supernatant but incorporated the 13C label after P. aeruginosa was cross-fed the R. mucilaginosa supernatant in minimal medium and artificial-sputum medium. We provide evidence that P. aeruginosa utilizes R. mucilaginosa-produced metabolites as precursors for generation of primary metabolites, including glutamate.IMPORTANCEPseudomonas aeruginosa is a dominant and persistent cystic fibrosis pathogen. Although P. aeruginosa is accompanied by other microbes in the airways of cystic fibrosis patients, few cystic fibrosis studies show how P. aeruginosa is affected by the metabolism of other bacteria. Here, we demonstrate that P. aeruginosa generates primary metabolites using substrates produced by another microbe that is prevalent in the airways of cystic fibrosis patients, Rothia mucilaginosa These results indicate that P. aeruginosa may get a metabolic boost from its microbial neighbor, which might contribute to its pathogenesis in the airways of cystic fibrosis patients.

Effect of ultrasound on the physicochemical and microbiological characteristics of Italian salami.

Italian salami were sonicated in different times (0, 3, 6 and 9min) using ultrasound bath (US, 25kHz). The effect of sonication on microbial growth (lactic acid bacteria and Micrococcaceae), lipid and protein oxidation, total heme pigments (THP), non heme iron (NHI) and metmyoglobin (MMb) was investigated during processing (0, 2, 15, and 28days) and storage (1, 30, and 120days). US enhanced growth of microorganisms (P<0.05), mainly for the treatment 9min of sonication. The lipid (peroxide value and TBARS) and protein (thiol group) oxidative reactions were accelerated by US (P<0.05) and they should be considered to maintain Italian salami quality. Sonication contributed to maintenance of THP (P<0.05), especially during storage. MMb pigment was not affected by sonication (P>0.05). This study presented some features of US application that could be explored in the manufacture of Italian salami.

Multiple plasmid-borne virulence genes of Clavibacter michiganensis ssp. capsici critical for disease development in pepper.

Clavibacter michiganensis ssp. capsici is a Gram-positive plant-pathogenic bacterium causing bacterial canker disease in pepper. Virulence genes and mechanisms of C. michiganensis ssp. capsici in pepper have not yet been studied. To identify virulence genes of C. michiganensis ssp. capsici, comparative genome analyses with C. michiganensis ssp. capsici and its related C. michiganensis subspecies, and functional analysis of its putative virulence genes during infection were performed. The C. michiganensis ssp. capsici type strain PF008 carries one chromosome (3.056 Mb) and two plasmids (39 kb pCM1Cmc and 145 kb pCM2Cmc ). The genome analyses showed that this bacterium lacks a chromosomal pathogenicity island and celA gene that are important for disease development by C. michiganensis ssp. michiganensis in tomato, but carries most putative virulence genes in both plasmids. Virulence of pCM1Cmc -cured C. michiganensis ssp. capsici was greatly reduced compared with the wild-type strain in pepper. The complementation analysis with pCM1Cmc -located putative virulence genes showed that at least five genes, chpE, chpG, ppaA1, ppaB1 and pelA1, encoding serine proteases or pectate lyase contribute to disease development in pepper. In conclusion, C. michiganensis ssp. capsici has a unique genome structure, and its multiple plasmid-borne genes play critical roles in virulence in pepper, either separately or together.

Biodegradation of atrazine by the novel Citricoccus sp. strain TT3.

A previously undescribed atrazine-degrading bacterial strain TT3 capable of growing with atrazine as its sole nitrogen source was isolated from soil at the wastewater outfall of a pesticide factory in China. Phenotypic characterization and 16S rRNA gene sequencing indicated that the isolate belonged to the genus Citricoccus. Polymerase chain reaction (PCR) analysis revealed that TT3 contained the atrazine-degrading genes trzN, atzB, and atzC. The range for growth and atrazine degradation of TT3 was found to be pH 6.0-11.0, with a preference for alkaline conditions. At 30°C and pH 7.0, the strain removed 50mg/L atrazine in 66h with 1% inoculum. These results demonstrate that Citricoccus sp. TT3 has great potential for bioremediation of atrazine-contaminated sites, particularly in alkaline environments. To the best of our knowledge, there are no previous reports of Citricoccus strains that degrade atrazine, and therefore this work provides a novel candidate for atrazine bioremediation.

Volatile organic compounds produced by a soil-isolate, Bacillus subtilis FA26 induce adverse ultra-structural changes to the cells of Clavibacter michiganensis ssp. sepedonicus, the causal agent of bacterial ring rot of potato.

Rhizobacterial volatile organic compounds (VOCs) play an important role in the suppression of soil-borne phytopathogens. In this study, the VOCs produced by a soil-isolate, Bacillus subtilis FA26, were evaluated in vitro for their antibacterial activity against Clavibacter michiganensis ssp. sepedonicus (Cms), the causal agent of bacterial ring rot of potato. The VOCs emitted by FA26 inhibited the growth of Cms significantly compared with the control. Scanning and transmission electron microscopy analyses revealed distorted colony morphology and a wide range of abnormalities in Cms cells exposed to the VOCs of FA26. Varying the inoculation strategy and inoculum size showed that the production and activity of the antibacterial VOCs of FA26 were dependent on the culture conditions. Headspace solid-phase microextraction/gas chromatography-mass spectrometry analyses revealed that FA26 produced 11 VOCs. Four VOCs (benzaldehyde, nonanal, benzothiazole and acetophenone) were associated with the antibacterial activity against Cms. The results suggested that the VOCs produced by FA26 could control the causal agent of bacterial ring rot of potato. This information will increase our understanding of the microbial interactions mediated by VOCs in nature and aid the development of safer strategies for controlling plant disease.

Isolation and identification methods of Rothia species in oral cavities.

Rothia dentocariosa and Rothia mucilaginosa which are Gram-positive bacteria are part of the normal flora in the human oral cavity and pharynx. Furthermore, Rothia aeria, which was first isolated from air samples in the Russian space station Mir, is predicted to be an oral inhabitant. Immunocompromised patients are often infected by these organisms, leading to various systemic diseases. The involvement of these organisms in oral infections has attracted little attention, and their distribution in the oral cavity has not been fully clarified because of difficulties in accurately identifying these organisms. A suitable selective medium for oral Rothia species, including R. aeria, is necessary to assess the veritable prevalence of these organisms in the oral cavity. To examine the bacterial population in the oral cavity, a novel selective medium (ORSM) was developed for isolating oral Rothia species in this study. ORSM consists of tryptone, sodium gluconate, Lab-Lemco powder, sodium fluoride, neutral acriflavin, lincomycin, colistin, and agar. The average growth recovery of oral Rothia species on ORSM was 96.7% compared with that on BHI-Y agar. Growth of other representative oral bacteria, i.e. genera Streptococcus, Actinomyces, Neisseria, and Corynebacterium, was remarkably inhibited on the selective medium. PCR primers were designed based on partial sequences of the 16S rDNA genes of oral Rothia species. These primers reacted to each organism and did not react to other non-oral Rothia species or representative oral bacteria. These results indicated that these primers are useful for identifying oral Rothia species. A simple multiplex PCR procedure using these primers was a reliable method of identifying oral Rothia species. The proportion of oral Rothia species in saliva samples collected from 20 subjects was examined by culture method using ORSM. Rothia dentocariosa, Rothia mucilaginosa, and R. aeria accounted for 1.3%, 5.9%, and 0.8% of the total cultivable bacteria number on BHI-Y agar in the oral cavities of all subjects, respectively. It was indicated that among oral Rothia species, R. mucilaginosa is most predominant in the oral cavity of humans. A novel selective medium, ORSM, was useful for isolating each oral Rothia species.

Modelling the growth kinetics of Kocuria marina DAGII as a function of single and binary substrate during batch production of ß-Cryptoxanthin.

In the present investigation, growth kinetics of Kocuria marina DAGII during batch production of ß-Cryptoxanthin (ß-CRX) was studied by considering the effect of glucose and maltose as a single and binary substrate. The importance of mixed substrate over single substrate has been emphasised in the present study. Different mathematical models namely, the Logistic model for cell growth, the Logistic mass balance equation for substrate consumption and the Luedeking-Piret model for ß-CRX production were successfully implemented. Model-based analyses for the single substrate experiments suggested that the concentrations of glucose and maltose higher than 7.5 and 10.0 g/L, respectively, inhibited the growth and ß-CRX production by K. marina DAGII. The Han and Levenspiel model and the Luong product inhibition model accurately described the cell growth in glucose and maltose substrate systems with a R 2 value of 0.9989 and 0.9998, respectively. The effect of glucose and maltose as binary substrate was further investigated. The binary substrate kinetics was well described using the sum-kinetics with interaction parameters model. The results of production kinetics revealed that the presence of binary substrate in the cultivation medium increased the biomass and ß-CRX yield significantly. This study is a first time detailed investigation on kinetic behaviours of K. marina DAGII during ß-CRX production. The parameters obtained in the study might be helpful for developing strategies for commercial production of ß-CRX by K. marina DAGII.

Characterization of specific spoilage organisms (SSOs) in vacuum-packed ham by culture-plating techniques and MiSeq next-generation sequencing technologies.

BACKGROUND: Knowledge regarding microaerophilic and anaerobic specific spoilage organisms (SSOs) is crucial for an appropriate evaluation of vacuum-packed ham. The objective of this study was to characterize the SSO community in vacuum-packed ham by a culture-dependent technique and MiSeq next-generation sequencing (NGS) platform. The relation between changes among the SSO group in the ham and changes in sensory characteristics of the product was also assessed. RESULTS: In the study, conventional microbiological analyses were employed in order to establish the participation of several groups of microorganisms in the deterioration of vacuum-packed ham. The diversity of the SSO group in the product was further assessed with the use of MiSeq NGS technology. The bacteria identified in sliced cooked ham belonged mostly to four phyla, namely Actinobacteria, Proteobacteria, Firmicutes and Bacteroidetes. A temperature of 4 °C favoured the development of mesophilic and psychrophilic/psychrotrophic flora, mainly Lactobacillaceae, Enterobacteriaceae and Micrococcaceae families. A high ratio of Brochothrix thermosphacta species and new, cold-tolerant Clostridium spp. was also observed. The growth of these microorganisms facilitated changes in the pH value and organoleptic characteristics of the product. CONCLUSION: This study confirms that the combination of culturing and MiSeq NGS technology improves the microbial evaluation of food. © 2016 Society of Chemical Industry.

PCR-Mediated Detection and Quantification of the Goss's Wilt Pathogen Clavibacter michiganensis subsp. nebraskensis Via a Novel Gene Target.

Goss's leaf blight and wilt of maize (corn) is a significant and reemerging disease caused by the bacterium Clavibacter michiganensis subsp. nebraskensis. Despite its importance, molecular tools for diagnosing and studying this disease remain limited. We report the identification of CMN_01184 as a novel gene target and its use in conventional PCR (cPCR) and SYBR green-based quantitative PCR (qPCR) assays for specific detection and quantification of C. michiganensis subsp. nebraskensis. The cPCR and qPCR assays based on primers targeting CMN_01184 specifically amplified only C. michiganensis subsp. nebraskensis among a diverse collection of 129 bacterial and fungal isolates, including multiple maize bacterial and fungal pathogens, environmental organisms from agricultural fields, and all known subspecies of C. michiganensis. Specificity of the assays for detection of only C. michiganensis subsp. nebraskensis was also validated with field samples of C. michiganensis subsp. nebraskensis-infected and uninfected maize leaves and C. michiganensis subsp. nebraskensis-infested and uninfested soil. Detection limits were determined at 30 and 3 ng of pure C. michiganensis subsp. nebraskensis DNA, and 100 and 10 CFU of C. michiganensis subsp. nebraskensis for the cPCR and qPCR assays, respectively. Infection of maize leaves by C. michiganensis subsp. nebraskensis was quantified from infected field samples and was standardized using an internal maize DNA control. These novel, specific, and sensitive PCR assays based on CMN_01184 are effective for diagnosis of Goss's wilt and for studies of the epidemiology and host-pathogen interactions of C. michiganensis subsp. nebraskensis.

Determination of factors responsible for the bioweathering of copper minerals from organic-rich copper-bearing Kupferschiefer black shale.

The aim of this study was to investigate the bioweathering of copper minerals present in the alkaline, copper-bearing and organic-rich Kupferschiefer black shale through the action of a consortium of indigenous lithobiontic, heterotrophic, neutrophilic bacteria isolated from this sedimentary rock. The involvement of microorganisms in the direct/enzymatic bioweathering of fossil organic matter of the rock was confirmed. As a result of bacterial activity, a spectrum of various organic compounds such as urea and phosphoric acid tributyl ester were released from the rock. These compounds indirectly act on the copper minerals occurring in the rock and cause them to weather. This process was reflected in the mobilization of copper, iron and sulfur and in changes in the appearance of copper minerals observed under reflected light. The potential role of identified enzymes in biodegradation of fossil organic matter and role of organic compounds released from black shale as a result of this process in copper minerals weathering was discussed. The presented results provide a new insight into the role of chemical compounds released by bacteria during fossil organic matter bioweathering potentially important in the cycling of copper and iron deposited in the sedimentary rock. The originality of the described phenomenon lies in the fact that the bioweathering of fossil organic matter and, consequently, of copper minerals occur simultaneously in the same environment, without any additional sources of energy, electrons and carbon.

Development of potentiometric equipment for the identification of altered dry-cured hams: A preliminary study.

Microbiological contamination in dry-cured ham can occur in the early stages of the process, a large number of microorganisms involved in spoilage can produce alterations in the product. These include non-common odours, which are detected at the end of the process by a procedure called "cala", consisting of a sharp instrument punctured in every ham; this is smelled by an expert taster, who classifies hams as good and altered hams. An electronic device would be suitable for this process given the large amount of hams. The present research aims to develop objective equipment based on the potentiometry technique that identifies altered hams. A probe was developed, containing silver, nickel and copper electrodes, and was employed to classify altered and unaltered hams prior to classification by a tester. The results shown lower Ag and higher Cu potential values for altered hams. The differences in potentiometric response reveal a classification model, although further studies are required to obtain a reliable classification model.