Geosmin Production Research Articles

Community of Environmental Streptomyces Related to Geosmin Development in Chinese Liquors

Diverse Streptomyces species act as geosmin producers in the Chinese liquor-making process. In this paper, the ecology of these Streptomyces species was analyzed using denaturing gradient gel electrophoresis (DGGE) of amplified Actinobacteria -specified rDNA. The result showed that Streptomyces were widely distributed during Daqu incubation, and multiple processing, geographic, and climate factors can affect their distribution and diversity. The genes associated with geosmin production were characterized in four geosmin-producing Streptomyces strains, all of which were isolated from geosmin-contaminated Daqu. On the basis of this information, a real-time PCR method was developed, enabling the detection of traces of Streptomyces in complex solid-state matrices. The primer was targeted at the gene coding for geosmin synthase (geoA). The real-time PCR method was found to be specific for geosmin-producing Streptomyces and did not show any cross-reactivity with geosmin-negative isolates, which are frequently present in the Chinese liquor-brewing process. Quantification of geoA in the Chinese liquor-making process could permit the monitoring of the level of geosmin producers prior to the occurrence of geosmin production. Comparison of the qPCR results based on the gene encoding geosmin synthase and Actinobacteria-specified rDNA showed that about 1-10% of the Actinobacteria carry the geosmin synthesis gene.

Impact of water quality on the bacterial populations and off-flavours in recirculating aquaculture systems

A variety of factors affecting water quality in recirculating aquaculture systems (RAS) are associated with the occurrence of off-flavours. In this study, we report the impact of water quality on the bacterial diversity and the occurrence of the geosmin-synthesis gene (geoA) in two RAS units operated for 252 days. Unit 2 displayed a higher level of turbidity and phosphate, which affected the fresh water quality compared with unit 1. In the biofilter, nitrification is one of the major processes by which high water quality is maintained. The bacterial population observed in the unit 1 biofilter was more stable throughout the experiment, with a higher level of nitrifying bacteria compared with the unit 2 biofilter. Geosmin appeared in fish flesh after 84 days in unit 2, whereas it appeared in unit 1 after 168 days, but at a much lower level. The geoA gene was detected in both units, 28 days prior to the detection of geosmin in fish flesh. In addition, we detected sequences associated with Sorangium and Nannocystis (Myxococcales): members of these genera are known to produce geosmin. These sequences were observed at an earlier time in unit 2 and at a higher level than in unit 1. This study confirms the advantages of new molecular methods to understand the occurrence of geosmin production in RAS.

Establishment of a real-time PCR method for quantification of geosmin-producing Streptomyces spp. in recirculating aquaculture systems

Geosmin and 2-methylisoborneol (MIB) have been associated with off-flavour problems in fish and seafood products, generating a strong negative impact for aquaculture industries. Although most of the producers of geosmin and MIB have been identified as Streptomyces species or cyanobacteria, Streptomyces spp. are thought to be responsible for the synthesis of these compounds in indoor recirculating aquaculture systems (RAS). The detection of genes involved in the synthesis of geosmin and MIB can be a relevant indicator of the beginning of off-flavour events in RAS. Here, we report a real-time polymerase chain reaction (qPCR) protocol targeting geoA sequences that encode a germacradienol synthase involved in geosmin synthesis. New geoA-related sequences were retrieved from eleven geosmin-producing Actinomycete strains, among them two Streptomyces strains isolated from two RAS. Combined with geoA-related sequences available in gene databases, we designed primers and standards suitable for qPCR assays targeting mainly Streptomyces geoA. Using our qPCR protocol, we succeeded in measuring the level of geoA copies in sand filter and biofilters in two RAS. This study is the first to apply qPCR assays to detect and quantify the geosmin synthesis gene ( geoA) in RAS. Quantification of geoA in RAS could permit the monitoring of the level of geosmin producers prior to the occurrence of geosmin production. This information will be most valuable for fish producers to manage further development of off-flavour events.

EXPRESSION OF THE GEOSMIN SYNTHASE GENE IN THE CYANOBACTERIUM ANABAENA CIRCINALIS AWQC318(1).

The occurrence of taste and odor episodes attributed to geosmin continues to trouble water utilities worldwide, and only recently have advances been made in our fundamental understanding of the biochemical and genetic mechanisms responsible for the production of geosmin in microorganisms. For the first time, we have examined the expression of the geosmin synthase gene and corresponding geosmin production by Anabaena circinalis Rabenh. ex Bornet et Flahault AWQC318 under conditions of continuous light illumination and the removal of light as a stimulus and demonstrate that the expression of geosmin synthase appears to be constitutive under these conditions. The decrease in geosmin synthase transcription post maximum cell numbers and stationary phase suggests that a decrease in isoprenoid synthesis may occur before a decrease in the transcription of ribosomal units as the process of cell death is initiated.

Development of phytoplankton communities and common off-flavors in a biofloc technology system used for the culture of channel catfish (Ictalurus punctatus)

The use of biofloc technology production systems continues to increase in the aquaculture industry worldwide. Recent research demonstrated that outdoor biofloc systems can be used to produce high yields of channel catfish (Ictalurus punctatus). However, studies have not yet been performed to determine the development and composition of phytoplankton communities and related off-flavor problems in these biofloc production systems. In this study, water samples were collected biweekly from May to November and channel catfish samples were collected during harvest in November from nine 18.6m2 biofloc culture tanks. Water and fillet samples were analyzed for levels of the common off-flavor compounds geosmin and 2-methylisoborneol (MIB). The development and composition of phytoplankton communities in each culture tank was also monitored. In addition, water and biofloc samples were evaluated to assess the microbial sources of geosmin and MIB within the culture tanks. Phytoplankton (including algae and cyanobacteria attached to bioflocs) biomass, as determined by concentrations of chlorophyll a in the water, gradually increased in all tanks over time. Phytoplankton communities that developed in the culture tanks were dominated by fast-growing, unicellular and small colonial types of green algae (chlorophytes) and diatoms (bacillariophytes) and slower growing, small colonial types of cyanobacteria (cyanophytes). A positive correlation (p<0.05) between cumulative feed addition and chlorophyll a concentration was found. Although geosmin and MIB were present in the culture water of each tank during most of the study, levels were typically low and only one tank yielded catfish with geosmin and MIB in their flesh at levels high enough to be designated as off-flavor. A positive correlation (p<0.05) between cumulative feed addition and MIB concentrations in the water of culture tanks indicates a greater potential for MIB-related off-flavor problems at high feed application rates. The microbial sources responsible for production of geosmin and MIB in the culture tanks remain unknown.

Geosmin causes off-flavour in arctic charr in recirculating aquaculture systems

The ‘earthy’ and ‘muddy’ off-flavours in pond-reared fish are due to the presence of geosmin or 2-methylisoborneol in the flesh of the fish. Similar off-flavours have been reported in fish raised in recirculating aquaculture systems (RAS); however, little information is available regarding the cause of these off-flavours. Our hypothesis was that earthy and muddy off-flavour compounds, found previously in pond-raised fish, are also responsible for off-flavours in fish raised in RAS. In this preliminary study, we examined water, biofilms in RAS and fillets from cultured arctic charr known to have off-flavours and requiring depuration using instrumental [solid-phase microextraction procedure and gas chromatograph-mass spectrometry (GC-MS)] and human sensory analyses. Geosmin was present in the samples taken from the biofilter and on the side walls of the tanks. Two-methylisoborneol was only found in low levels in the samples. The GC-MS results indicated the presence of geosmin in the fillets (705 ng kg−1), but lower levels were found in the water (30.5 ng L−1). Sensory analyses also detected an earthy flavour (i.e., geosmin presence) in the fillets, and, therefore, it appears that geosmin is the main compound responsible for the off-flavour in RAS. Further studies are being performed to identify the microorganisms responsible for geosmin production in RAS.

Cyanobacterial population and harmful metabolites dynamics during a bloom in Yanghe Reservoir, North China

A significant outbreak of odorous and toxic cyanobacteria occurred in Yanghe Reservoir, North China, in the summer of 2007. The dominant species was Anabaena spiroides and it was accompanied with the occurrence of very high concentrations of the odor metabolite geosmin. The event included two cycles of growth of A. spiroides: the first one of approximately 16 days between 21 June and 8 July, with the peak density 70,000 cells mL −1 on 2 July, and the second smaller bloom of over 50 days between 9 July and the end of August. The bloom also included a range of species of Microcystis, predominantly M. aeruginosa, as the second largest population, which varied from 5800 cells mL −1 to 28,000 cells mL −1 in the first cycle and from 2000 cells mL −1 to 11,000 cells mL −1 in the second. Geosmin reached the peak value of 7100 ng L −1 on 3 July, which was the highest value ever reported. The average geosmin production potential for A. spiroides cells was approximately 0.1 pg cell −1, which was also very high and 85–97% of geosmin was found within the cells and in addition the dissolved geosmin did not increase even during the cyanobacterial decay period. In addition to odor metabolites, three microcystin variants MC-LR, MC-RR, MC-YR and anatoxin-a, most of which were intracellular, were detected, with MC-RR being the dominant cyanotoxin. The highest cyanotoxin concentrations were: dissolved: MC-RR 1.56 μg L −1, MC-YR 0.066 μg L −1, MC-LR 0.544 μg L −1, anatoxin-a 0.106 μg L −1; intracellular: MC-RR 70.1 μg L −1, MC-YR 3.76 μg L −1, MC-LR 24.6 μg L −1, anatoxin-a 0.184 μg L −1, and these occurred on 2 July. The correlation between geosmin and A. spiroides was excellent ( R 2 = 0.912), however the correlation between anatoxin-a concentrations and Anabaena densities and between MCs concentrations and Microcystis densities was not as strong. This study demonstrated the relatively complex requirement for monitoring cyanobacterial blooms in this lake in China, where both the range of odors and cyanotoxins can be produced in concentrations which can change rapidly over a short time along with the bloom composition. It also demonstrated that both odor compounds and cyanotoxins should be considered in terms of the potential hazard to public water supply when a bloom was dominated by Anabaena and Microcystis occurs.

Isolation, identification and odour-producing abilities of geosmin/2-MIB in actinomycetes from sediments in Lake Lotus, China

This study aimed to determine whether actinomycetes could contribute to the odours occurring in Lake Lotus, China. In total, 55 actinomycete strains were isolated from sediments in Lake Lotus and their odorous metabolites, geosmin and 2-methylisoborneol (2-MIB), were identified using HSPME-GC-MS. Results showed that 24, 23 and 34 strains produced geosmin and/or 2-MIB in Gause, TSB and M liquid media, respectively. Of odour-producing actinomycetes, most could produce geosmin and some produced both metabolites, while few of them produced only 2-MIB. Six isolates with high-level odour were selected for further investigation. Their biomass and odour-producing abilities were monitored in both the slants and liquid media. The results suggest that TSB was the most suitable medium for the growth of mycelium; Gause and M slant supported good production of spores, while M liquid medium was the most favourable for the production of both geosmin and 2-MIB. Those strains that produced geosmin only were less influenced than those that produced both geosmin and 2-MIB under shaking conditions. The results indicate that actinomycetes from sediments should be taken into consideration when off-flavours occur in water columns. According to the 16S rRNA sequences, six actinomycetes were classified in the Streptomyces.

Occurrence of odour-causing compounds in different source waters of China

In this study, occurrence of the main odorous compounds in different types of source water in China, including rivers, lakes and reservoirs, were investigated by integrating the methods of flavour profile analysis (FPA) and sensory-GC/MS analysis. Simultaneous distillation extraction (SDE) was chosen as the preconcentration technique while the odour-causing compounds were identified with sensory-GC/MS. It was found that MIB, which is mainly produced by some cyanobacteria and actinomycetes, was one of the most widely distributed odour-causing compounds, regardless of source type. A peak MIB concentration of approximately 170 ng l(-1) was recorded in MY Reservoir in October 2005. Among the nine investigated sites, YH Reservoir in Hebei Province was the only site showing geosmin production. A peak geosmin concentration exceeding 7,000 ng l(-1) was detected in July 2007. In June 2007, a serious taste/odour (T/O) event occurred in Wuxi City, and the offensive flavour was a strong septic or marshy odour caused by dimethyl trisulfide (431 ng l(-1) in finished water). Septic/swampy and earthy/musty odours occurred in source waters of Shanghai and Guangzhou. It was found that dimethyl trisulfide was mainly responsible for the septic/swampy odour and MIB for the earthy/musty one. Dimethyl trisulfide might be an important T/O-causing compound in China.

Exploration of geosmin synthase from Streptomyces peucetius ATCC 27952 by deletion of doxorubicin biosynthetic gene cluster

Thorough investigation of Streptomyces peucetius ATCC 27952 genome revealed a sesquiterpene synthase, named spterp13, which encodes a putative protein of 732 amino acids with significant similarity to S. avermitilis MA-4680 (SAV2163, GeoA) and S. coelicolor A3(2) (SCO6073). The proteins encoded by SAV2163 and SCO6073 produce geosmin in the respective strains. However, the spterp13 gene seemed to be silent in S. peucetius. Deletion of the doxorubicin gene cluster from S. peucetius resulted in increased cell growth rate along with detectable production of geosmin. When we over expressed the spterp13 gene in S. peucetius DM07 under the control of an ermE* promoter, 2.4 +/- 0.4-fold enhanced production of geosmin was observed.

Development of predictive models for geosmin-related taste and odor in Kansas, USA, drinking water reservoirs

The presence of taste and odor compounds can greatly reduce the quality of drinking water supplies. Because the monetary costs associated with the removal of these compounds can be high, it is impractical for most facilities to continuously treat their raw water. Instead, new tools are needed to help predict when taste and odor events may be most likely to occur. Water quality data were collected between June and October in 2006–2007 from five Kansas (USA) reservoirs in order to develop predictive models for geosmin, a major taste and odor compound; two of these reservoirs were also sampled during specific taste and odor events in December 2006 and January 2007. Lake trophic state alone was not a good predictor of geosmin concentrations as the highest average geosmin concentration was observed in the reservoir with the lowest nutrient and chlorophyll a concentrations. In addition, taste and odor events were not confined to summer months; elevated geosmin concentrations were observed in several reservoirs during the winter. Growth limitation by inorganic phosphorus appeared to be the primary determinant of geosmin production by algal cells in these reservoirs.

Reservoir Circulation Reduces Geosmin Production

This article discusses an effort at Wake Forest Reservoir, North Carolina, to mitigate taste and odor issues caused by microorganisms. A new approach was used that consisted of releasing compressed air into the bottom of the reservoir through an air‐diffuser array. To study the link between water chemistry, stratification, and taste and odor production, limnocorrals – circular enclosures that isolate portions of the water column from the rest of the impoundment in which various treatments can be tested – were installed to test the potential effectiveness of artificial destratification and subsurface copper sulfate injection. Geosmin production was lower in the subsurface copper sulfate and aerated limnocorrals compared to the controls. Bolstered by reduced geosmin, manganese, and iron in the limnocorral treatments, the town of Wake Forest continued to apply copper sulfate to the surface waters of the impoundment and began to destratify the water column by centering a diffuser array on top of the deep basin. Information is presented from deep basin limnology studies that continued over a two year period, including the following: temperature variations; dissolved oxygen; copper sulfate application; and, taste‐ and odor‐producing microorganisms.

Isolation and Characterization of the Gene Associated with Geosmin Production in Cyanobacteria

Geosmin is a secondary metabolite responsible for earthy tastes and odors in potable water supplies. Geosmin continues to be a challenge to water utility management regimes and remains one of the most common causes of consumer complaints, as the taste of "dirty" water may suggest a failed disinfection regime and that the water may be unsafe to drink. Although cyanobacteria have been reported to be largely responsible for these taste and odor events, the answer as to how or why geosmin is produced has eluded researchers. We describe here for the first time the mechanism by which geosmin is produced in a model cyanobacterium, Nostoc punctiforme PCC 73102 (ATCC 29133), which we demonstrate utilizes a single enzyme to catalyze the cyclization of farnesyl diphosphate to geosmin. Using this information, we have developed a PCR-based assay that allows the rapid detection of geosmin-producing cyanobacteria. This test may be utilized to confirm and track the emergence of taste and odor-producing cyanobacteria in any given water body and thus can be used as an early warning system by managers of water bodies that may suffer from adverse taste and odor episodes.

Effects of temperature and light on the growth and geosmin production of Lyngbya kuetzingii (Cyanophyta)

The effects of temperature and light on the growth and geosmin production of Lyngbya kuetzingii were determined. Of the three temperatures tested, 10, 25 and 35°C, the maximal geosmin concentration and geosmin productivity were yielded at 10°C, while the highest chl a production was observed at 25°C. In the studies on light intensity, the maximal geosmin concentration and geosmin productivity were observed at 10 μmol m−2 s−1, while the highest chl a production was at 20 μmol m−2 s−1. It was suggested that more geosmin was synthesized with lower chl a demand. Meanwhile, the relative amounts of extra- and intracellular geosmin were investigated. Under optimum growth conditions (20 μmol m−2 s−1, 25°C; BG-11 medium), the amounts of extracellular geosmin increased as the growth progressed and reached the maximum in the stationary phase, while the intracellular geosmin reached its maximum value in the late exponential phase, and then began to decline. However, under the low temperature (10°C) or light (10 μmol m−2 s−1) conditions, more intracellular geosmin was synthesized and mainly accumulated in the cells. The proportions of extracellular geosmin were high, to 33.33 and 32.27%, respectively, during the stationary phase at 35°C and 20 μmol m−2 s−1. It was indicated that low temperature or light could stimulate geosmin production and favor the accumulation of geosmin in cells, while more intracellular geosmin may be released into the medium at higher temperatures or optimum light intensity.

Identification of conditions underlying production of geosmin and 2-methylisoborneol in a recirculating system

Geosmin and 2-methylisoborneol (MIB) are semi-volatile terpenoid compounds produced as secondary metabolites by benthic and planktonic cyanobacteria, several genera of fungi, and various actinomycetes. These off-flavor compounds pose a heavy economic burden in the aquaculture industry rendering fish unmarketable unless purified by purging with large quantities of clean water. In the present study, the presence of off-flavor compounds was examined in a recirculating aquaculture system (RAS) for tilapia culture. In this zero-discharge system, where water from the fish basins is recirculated through parallel aerobic (drum filter and a trickling filter) and anaerobic treatment loops (sedimentation/digestion basin), concentrations of geosmin and, in particular, MIB were highest in the aerobic treatment loop. Lowest concentrations were detected in the anaerobic treatment loop. This latter finding pointed toward a possible reduction of these compounds in this basin. Two bacterial strains of the streptomycetes family were isolated from the aerobic, organic-rich, drum filter and the nitrifying trickling filter. In vitro tests with these isolates, closely related to Streptomyces roseoflavus and Streptomyces thermocarboxydus, revealed that MIB production exceeded geosmin production under all conditions tested and was significantly higher under aerobic than under anoxic conditions. Under the latter conditions, with nitrate as an electron donor, the S. roseoflavus-like isolate was capable of denitrification. Based on the results obtained in this study, it was concluded that aerobic, organic-rich conditions stimulate the growth of actinomycetes and subsequent production of geosmin and MIB in the system. The observed reduction of these compounds in the anaerobic water treatment component may serve in designing treatment steps aimed at alleviating the problem of geosmin and MIB accumulation in recirculating systems.

Impact of the Botrytis cinerea strain and metabolism on (−)-geosmin production by Penicillium expansum in grape juice

Geosmin, an off-flavour of some rotten grapes, has been implicated in wine defects. Botrytis cinerea and Penicillium expansum were the most common among the numerous microorganisms isolated from rotten grapes. P. expansum produces geosmin on model media but not healthy grape juice. However, geosmin synthesis by P. expansum was demonstrated in grape juice and on crushed grapes that had been pre-cultured with certain B. cinerea strains. 34 out of 156 B. cinerea strains ([bot +] phenotype) isolated from the centre of grape bunches were able to induce high geosmin production, up to 494 ng/l, by P. expansum in grape juice. A study of the impact of grape juice composition on geosmin synthesis by P. expansum revealed the importance of nitrogen composition, particularly amino-acid deficiency. Metabolism of amino acids by B. cinerea was shown to be favourable to geosmin synthesis by P. expansum. However, the amino-acid and ammonium concentrations in grape juices pre-cultured with B. cinerea [bot -] and [bot +] strains were very similar implying that other factors are involved as well. Indeed, an ethanol-precipitable fraction, probably a polysaccharide, synthesized by B. cinerea [bot -], but not [bot +] strains, inhibited geosmin production by P. expansum.

A comparison of biofilms from macrophytes and rocks for taste and odour producers in the St. Lawrence River

Given their widespread and prolific annual development in the St. Lawrence River (SLR), macrophytes (i.e. submerged aquatic plants) represent large surface areas for biofilm growth and potentially important sites for associated production of taste and odour (T&O) compounds. We therefore evaluated the importance of submerged macrophytes and their associated biofilms for production of T&O compounds, 2-methylisoborneol (MIB) and geosmin (GM), compared with biofilms from adjacent rocks. We also tested the hypothesis that production of these compounds would differ between macrophyte species, based on the premise that they are not inert substrates but directly influence the communities that colonise their surfaces. Samples collected from transects across the SLR between Kingston and Cornwall, ON were dominated by the flat-bladed Vallisneria spp., and the leafed Myriophyllum spicatum, Elodea canadensis, Chara spp., Potamgeton spp., and Ceratophyllum spp. Overall, MIB and GM levels in biofilms ranged widely between samples. Expressed per g dry weight of biofilm, median levels from macrophyte were 50 (range 1-5000) ng MIB g(-1) and 10 (<1 to 580) ng GM g(-1) compared with 50 (range 5-970) ng MIB g(-1) and 160 (1-1600) ng GM g(-1) from rocks. Based on non-parametric statistical analysis, levels of GM were higher on a g dry weight basis in biofilms from rocks than macrophytes (P = 0.02), but MIB levels were similar (P = 0.94). However, when normalised for differences in substrate surface area (i.e. ng cm(-2)), levels of both MIB and GM were higher in biofilms from rocks than from macrophytes (P < 0.01). There were no discernable differences in MIB and GM concentrations from biofilms of different macrophytes based on either g dry weight sample or surface area (P > 0.05). Overlying water (OLW) concentrations ranged between 2-45 ng L(-1) for MIB and 5-30 ng L(-1) for GM and were not correlated with levels in adjacent biofilms. However, OLW concentrations peaked in shallow, low energy embayments consistent with enhanced production and release of MIB and GM in nearshore areas. The results support our previous work showing the importance of biofilms on various surfaces (rocks, macrophytes and zebra mussels) for MIB and GM production in the SLR, but suggest that inert surfaces like rocks are more productive sites per unit surface area than macrophytes.

Relationship between volatile odorous substances and production of avermectins byStreptomyces avermitilis

The time course of production of odorous compounds, i.e. geosmin and oxolones, and of avermectins was determined during the cultivation of S. avermitilis in flasks, 1- and 50-L fermentors. The amount of the antibiotics increased with increasing cultivation time up to more than 2 g/L while the concentration of geosmin rose to more than 4 mg/L. Cultivation without reflux condenser resulted in a lower product formation due to the higher stripping of geosmin. A relatively tight correlation was found between the production of geosmin and the production of avermectins. The production of oxolones peaked on cultivation days 3-5, the sum of oxolones being 60 microg/L. Subsequently, the production dropped below a measurable level. This can be explained as being due to the inhibition of oxolone production by geosmin.

A CAPS test allowing a rapid distinction of Penicillium expansum among fungal species collected on grape berries, inferred from the sequence and secondary structure of the mitochondrial SSU-rRNA

Penicillium expansum is a fungal species highly damageable for the postharvest conservation of numerous fruits. In vineyards, this fungus is sometimes isolated from grape berries where its presence may lead to the production of geosmin, a powerful earthy odorant, which can impair grapes and wines aromas. However, the discrimination of P. expansum from related fungi is difficult because it is based on ambiguous phenotypic characters and/or expensive and time-consuming molecular tests. In this context, the complete sequences and secondary structures of Penicillium expansum and Penicillium thomii mitochondrial SSU-rRNAs were achieved and compared with those of two other phylogenetically related Ascomycota: Penicillium chrysogenum and Emericella nidulans. The comparison has shown a high conservation in size and sequence of the core and of the variable domains (more than 80% of nt identity) of the four SSU-rRNAs, arguing for a close phylogenetic relationship between these four species of the Trichocomaceae family. Large (from 10 to 18 nt) inserted/deleted (indel) sequences were evidenced in the V1, V5 and V6 variable domains. The size variations (10 to 18 nt) of the V1 indel sequence allowed the distinction of the four species; the V5 indel (15 nt) was specifically recovered in E. nidulans; the V6 indel (16 nt), shared by the three Penicillium species, was lacking in E. nidulans. A couple of conserved primers (UI/R2) were defined to generate a PCR product containing the V1 to V5 variable domains. This product contained the two regions of the four SSU-rRNAs showing the highest rates of nt substitutions, namely the V2 variable domain and, surprisingly, a helix (H17) of the core. The H17 sequence was shown to specifically possess in P. expansum a recognition site for the ClaI restriction endonuclease. Hence, this enzyme generates a digestion pattern of the PCR product with two bands (350 bp + 500 bp), specific to P. expansum and easily separable by agarose gel electrophoresis. This leads to a CAPS test usable for P. expansum discrimination among grape berries isolated filamentous fungi. The CAPS test was validated by a comparative analysis involving 29 strains belonging to 17 species currently isolated from grape berries in the Bordeaux vineyards.

Potential of an electronic nose for the early detection and differentiation between Streptomyces in potable water

Much attention has been focused on the production of musty-aroma compounds such as geosmin and their impact on the quality of fresh water and water-cultured raised fish and sea-food. At present, there are no efficient means of removing these off-flavours from water. Therefore, the rapid detection of geosmin-producing microorganisms, in particular the genus Streptomyces, at early stages of differentiation, is still the best option in preventing deterioration of water quality. We investigated the potential of an electronic nose consisting of an array of 14 conducting polymer sensors for the rapid and early detection of Streptomyces spores in reverse osmosis and tap water. Geosmin solutions in water at different concentrations were also prepared for headspace analysis in order to study the e-nose potential for detecting geosmin production in this environment. Normalised divergence data were analysed using principal component analysis (PCA) and discriminant function analysis (DFA). Data indicates that an e-nose could be employed to detect and monitor early activity of Streptomyces in water at different stages of differentiation, as well as to discriminate between different species based on their volatile production patterns. It also suggests that it could be used for monitoring geosmin production in water and possibly set threshold odour levels, as a routine task for specific water-screening purposes.