Genetic diversity and structure of the coffee leaf rust fungus Hemileia vastatrix across different coffee management systems in Ethiopia / Diversidad genética y estructura del hongo de la roya del café Hemileia vastatrix en diferentes sistemas de manejo del café en Etiopía

Although coffee leaf rust (CLR), caused by Hemileia vastatrix, poses an increasing threat to coffee production in Ethiopia, little is known regarding its genetic diversity and structure and how these are affected by coffee management. Here, we used genetic fingerprinting based on sequence-related amplified polymorphism (SRAP) markers to genotype H. vastatrix samples from different coffee shrubs, across 40 sites, covering four coffee production systems (forest coffee, semi plantation coffee, home garden coffee, and plantation coffee) and different altitudes in Ethiopia. In total, 96 H. vastatrix samples were successfully genotyped with three primer combinations, producing a total of 79 scorable bands. We found 35.44% of amplified bands to be polymorphic, and the polymorphic information content (PIC) was 0.45, suggesting high genetic diversity among our CLR isolates. We also found significant isolation-by-distance across the samples investigated and detected significant differences in fungal genetic composition among plantation coffee and home garden coffee and a marginally significant difference among plantation coffee and forest coffee. Furthermore, we found a significant effect of altitude on CLR genetic composition in the forest coffee and plantation systems. Our results suggest that both spore dispersal and different selection pressures in the different coffee management systems are likely responsible for the observed high genetic diversity and genetic structure of CLR isolates in Ethiopia. When selecting Ethiopian coffee genotypes for crop improvement, it is important that these genotypes carry some resistance against CLR. Because our study shows large variation in genetic composition across relatively short geographical distances, a broad selection of rust isolates must be used for coffee resistance screening.(AU)

Parallel host shifts in a bacterial plant pathogen suggest independent genetic solutions.

While there are documented host shifts in many bacterial plant pathogens, the genetic foundation of host shifts is largely unknown. Xylella fastidiosa is a bacterial pathogen found in over 600 host plant species. Two parallel host shifts occurred-in Brazil and Italy-in which X. fastidiosa adapted to infect olive trees, whereas related strains infected coffee. Using 10 novel whole-genome sequences from an olive-infecting population in Brazil, we investigated whether these olive-infecting strains diverged from closely related coffee-infecting strains. Several single-nucleotide polymorphisms, many derived from recombination events, and gene gain and loss events separated olive-infecting strains from coffee-infecting strains in this clade. The olive-specific variation suggests that this event was a host jump with genetic isolation between coffee- and olive-infecting X. fastidiosa populations. Next, we investigated the hypothesis of genetic convergence in the host shift from coffee to olive in both populations (Brazil and Italy). Each clade had multiple mutations and gene gain and loss events unique to olive, yet no overlap between clades. Using a genome-wide association study technique, we did not find any plausible candidates for convergence. Overall, this work suggests that the two populations adapted to infect olive trees through independent genetic solutions.

The essential role of spontaneous and starter yeasts in cocoa and coffee fermentation.

Yeasts are important microorganisms used in different fermentation processes. The cocoa beans must go through a correct fermentation process to obtain good-quality chocolate, which involves the action of yeasts and bacteria, and yeasts play a crucial role since they act in the first days of fermentation. In coffee, several studies have shown that the microbiota in the fruits is also a relevant factor. The fermentation process (regardless of the processing type) improves the beverage's quality. In this sense, studies using starter cultures in these two raw materials are important for better control of the process, and optimization of fermentation time, in addition to the improvement and diversification of volatile and non-volatile compounds produced by yeasts. Thus, this review discusses the importance and role of yeasts during fermentation, their metabolism, the produced compounds, and how yeast and the different chemical reactions help increase the quality of chocolate and coffee.

Inoculation of yeast and bacterium in wet-processed Coffea canephora.

This study evaluated the inoculation of Meyerozyma guilliermondii and Bacillus licheniformis, separately or in co-culture, in wet-processed conilon coffee. Wet fermentation was conducted for 48 h. Mesophilic bacteria, lactic acid bacteria, yeasts, and filamentous fungi were counted during fermentation. The inoculation of B. licheniformis and M. guilliermondii stimulated the multiplication of lactic acid bacteria. Acetic, citric, lactic, oxalic, malic, succinic, tartaric acids, glucose, and fructose were identified in all treatments at different concentrations. Methyl salicylate, 2-heptanol, 2-nonanol, and heptanone were found during fermentation. Methylpyrazine, 2,6-dimethylpyrazine, 2,5-dimethylpyrazine, and 3-ethyl-2,5-dimethylpyrazine identified after roasting assigned notes of "almond" and "chocolate" to the beverages. All treatments were classified as "premium," with the B. licheniformis treatment receiving the highest score. Bacillus licheniformis obtained better performance in fermentation, increasing coffee score and producing volatile compounds that provided positive sensory notes to the beverage.

Mathematical modelling and analysis of coffee berry disease dynamics on a coffee farm.

This paper focuses on a mathematical model for coffee berry disease infestation dynamics. This model considers coffee berry and vector populations with the interaction of fungal pathogens. In order to gain an insight into the global dynamics of coffee berry disease transmission and eradication on any given coffee farm, the assumption of logistic growth with a carrying capacity reflects the fact that the amount of coffee plants depends on the limited size of the coffee farm. First, we show that all solutions of the chosen model are bounded and non-negative with positive initial data in a feasible region. Subsequently, endemic and disease-free equilibrium points are calculated. The basic reproduction number with respect to the coffee berry disease-free equilibrium point is derived using a next generation matrix approach. Furthermore, the local stability of the equilibria is established based on the Jacobian matrix and Routh Hurwitz criteria. The global stability of the equilibria is also proved by using the Lyapunov function. Moreover, bifurcation analysis is proved by the center manifold theory. The sensitivity indices for the basic reproduction number with respect to the main parameters are determined. Finally, the numerical simulations show the agreement with the analytical results of the model analysis.

Coinoculation of lactic acid bacteria and yeasts increases the quality of wet fermented Arabica coffee.

Wet coffee fermentation is widely used in coffee-producing regions such as Colombia and Hawaii, but it is not widespread in Brazil. This study aimed to evaluate inoculating the lactic acid bacteria Leuconostoc mesenteroides CCMA1105 and Lactiplantibacillus plantarum CCMA 1065 and the yeasts Saccharomyces cerevisiae CCMA0543 and Torulaspora delbrueckii CCMA0684 as starter cultures on wet coffee fermentation using the SIAF method (self-induced anaerobiosis fermentation). The microbial activity resulted in high consumption of the carbohydrates glucose (98.6%), fructose (97.6%), and sucrose (100%), in addition to the production of lactic and acetic acids, impacting the final quality of the beverage. A total of 108 volatile compounds belonging to 17 classes were identified in the green and roasted coffee samples, including 2,3-butanediol produced by lactic acid bacteria, contributing to coffee's aromatic profile. The final scores for the coffees from the different fermentations ranged from 79.0 to 83.25. The inoculated fermentations were classified as specialty according to the Specialty Coffee Association. Therefore, whole coffee fruit processed via wet using SIAF method and yeast and lactic acid bacteria starter is an alternative for improving wet fermented coffee quality and obtaining coffee beverages with a different sensory profile.

Constitutive Defense Strategy of Coffee Under Field Conditions: A Comparative Assessment of Resistant and Susceptible Cultivars to Rust.

Coffea arabica is the most economically important coffee species worldwide. However, its production is severely limited by diseases such as rust. The mechanisms underlying constitutive defense responses in coffee are still poorly understood, compared with induced defense mechanisms. We aimed to characterize constitutive defense responses of thirteen cultivars of C. arabica. Cultivars were classified under field conditions according to the level of resistance to rust: resistant (R), moderately resistant (MR), and susceptible (S). Based on this classification, the stability of eight reference genes (RGs) was evaluated. The most stable RGs were EF1α, APT1, and 24S. We also evaluated the expression of CaWRKY1, CaPAL1, CaCAD1, and CaPOX1, and activities of PAL, CAD, and POX, which are involved in lignin biosynthesis, and leaf content of total phenolic compounds and lignin. Gene expression and enzymatic activity were not correlated with defense metabolites in the R cultivar group but showed a negative correlation with phenolic compounds in MR cultivars. Cultivar S showed positive correlations of gene expression and enzyme activity with phenolic compounds. These results may assist coffee breeding programs regarding selection of genotypes and in optimization of rust resistance.

Evaluation of fungal contamination and ochratoxin A detection in different types of coffee by HPLC-based method.

BACKGROUND: Mycotoxins are secondary fungal metabolites that are produced by some toxigenic fungi on foodstuffs which are poisoning and potentiate for human's health hazards. In coffee samples, ochratoxin A and fungal contamination were examined. METHODS: Immunoaffinity columns were used for treating of all 50 samples from four types of coffee, after that high-performance liquid chromatography was used for determining the amount of ochratoxin. For the identification of fungi, all coffee samples were cultured in appropriated media. RESULTS: The results showed that all samples were contaminated by ochratoxin A but only up to 50% of them had toxins higher than acceptable level as detected in black beans (47%), green beans (33.3%), torch (33.3%), and espresso (25%). Black coffee had a higher mean concentration of ochratoxin A than green coffee. CONCLUSION: Predominant fungi isolated from coffee samples were Aspergillus species. Finally, careful monitoring of mycotoxins in coffee samples is essential to improve the quality of this favorable beverage in future.

Epidemics and the future of coffee production.

In this perspective, we draw on recent scientific research on the coffee leaf rust (CLR) epidemic that severely impacted several countries across Latin America and the Caribbean over the last decade, to explore how the socioeconomic impacts from COVID-19 could lead to the reemergence of another rust epidemic. We describe how past CLR outbreaks have been linked to reduced crop care and investment in coffee farms, as evidenced in the years following the 2008 global financial crisis. We discuss relationships between CLR incidence, farmer-scale agricultural practices, and economic signals transferred through global and local effects. We contextualize how current COVID-19 impacts on labor, unemployment, stay-at-home orders, and international border policies could affect farmer investments in coffee plants and in turn create conditions favorable for future shocks. We conclude by arguing that COVID-19's socioeconomic disruptions are likely to drive the coffee industry into another severe production crisis. While this argument illustrates the vulnerabilities that come from a globalized coffee system, it also highlights the necessity of ensuring the well-being of all. By increasing investments in coffee institutions and paying smallholders more, we can create a fairer and healthier system that is more resilient to future social-ecological shocks.

Growth, survival, and metabolic activities of probiotics Lactobacillus rhamnosus GG and Saccharomyces cerevisiae var. boulardii CNCM-I745 in fermented coffee brews.

Amidst rising demand for non-dairy probiotic foods, and growing interest in coffees with added functionalities, it would be opportune to ferment coffee brews with probiotics. However, challenges exist in maintaining probiotic viability in high-moisture food products. Here, we aimed to enhance the viability of the probiotic bacteria, Lactobacillus rhamnosus GG, in coffee brews by co-culturing with the probiotic yeast, Saccharomyces cerevisiae var. boulardii CNCM-I745. The yeast significantly enhanced the viability of L. rhamnosus GG, as bacterial populations beyond 7 Log CFU/mL were maintained throughout 14 weeks of storage at 4 and 25 °C. In contrast, the single culture of L. rhamnosus GG suffered viability losses below 6 Log CFU/mL within 10 weeks at 4 °C, and 3 weeks at 25 °C. Growth and survival of S. boulardii CNCM-I745 remained unaffected by the presence of L. rhamnosus GG. Volatile profiles of coffee brews were altered by probiotic metabolic activities, but co-culturing led to suppressed generation of diacetyl and ethanol compared to single cultures. Probiotic fermentation did not alter principal coffee bioactive compounds and antioxidant capacities; however, declines in peroxyl radical scavenging capacities were observed after ambient storage. Overall, we illustrate that yeasts are effective in enhancing probiotic bacterial viability in coffee brews, which may be useful in developing shelf stable probiotic food products.

Microbial diversity and chemical characteristics of Coffea canephora grown in different environments and processed by dry method.

This study aimed to assess the microbial diversity in Coffea canephora grown in four different environments of Espirito Santo state, Brazil. Coffee cherries of two different altitudes (300 and 600 m) and two terrain aspects (Southeast-facing and Northwest-facing slopes) were processed by the dry method. Samples were collected during the drying/fermentation process. Microorganisms were counted, isolated, and identified by MALDI-TOF, followed by sequencing of the ribosomal region. Sugars and organic acids were quantified by HPLC and volatile compounds of the roasted coffees were evaluated by GC-MS. Bacteria population presented a significant number of isolates as well as higher counts during the drying/fermentation process with respect to the population of yeasts. The principal genera of microorganisms found were Bacillus, Pichia, Candida, and Meyerozyma. Meyerozyma guilliermondii was the most frequent yeast in all environments. On the other hand, Pichia kluyveri was found only in coffee cherries from the 600 m altitude. The highest concentration of acetic and succinic acids observed was 6.06 mg/g and 0.84 mg/g, respectively. Sucrose concentrations ranged from 0.68 to 5.30 mg/g, fructose from 1.30 to 4.60 mg/g, and glucose from 0.24 to 1.25 mg/g. Thirty-six volatile compounds, belonging to the groups of pyrazines, alcohols, aldehydes, ketones, and furans were identified in roasted coffee, with differences between altitude and terrain aspects. Information about microbial diversity is crucial to better understand the coffee quality and distinct characteristics of coffee produced in different environments.

Isolation of microorganisms from the feces of ring-tailed coati related to the production of "misha coffee" in the central forest of Peru and evaluation of some features of technological importance.

A diversity of yeasts and lactic bacteria were isolated from the feces of ring-tailed coaties bred in captivity and related to the production of "misha coffee". Isolation of yeasts was carried out using oxytetracycline-glucose-yeast extract agar containing 100 mg/L oxytetracycline and, lactic bacteria using de Man-Rogosa and Sharpe agar containing 20 mg/L of vancomicin. Then, isolates were biochemically analysed using API strips (ID 32C for yeasts and 50CHL for lactic bacteria) followed by 16S and 26S rRNA gene sequencing. Among the yeasts, Debaryomyces hansenii, Pichia kluyveri, Pichia kudriavzevii, and Candida sorboxilosa were the most frequent, whereas Weissella cibaria, Weissella paramesenteroides, Enterococcus thailandicus and Enterococcus faecalis were the most important lactic bacteria. Cultivation of the isolated yeasts under agitated conditions, showed that Pichia kluyveri LBFT.Lev3 (0.15 ± 0.01 h-1) and Pichia kudriavzevii LBTF.Lev7 (0.14 ± 0.01 h-1) had higher specific growth rates than Debaryomyces hansenii LBFT.Lev9 (0.09 ± 0.01 h-1), whereas cultivation of lactic bacteria under static fashion showed that Weisella paramesenteroides LBTF.Bal2 (0.16 ± 0.01 h-1) and Weisella cibaria LBTF.Bal3 (0.18 ± 0.01 h-1) had better growth than Enterococcus thailandicus LBTF.Bal1 (0.1 ± 0.015 h-1) and Enterococcus faecalis LBTF.Bal7 (0.14 ± 0.01 h-1). Additionally, evaluation of pectinolytic activity revealed that Pichia kudriavzevii LBTF.Lev7 and Debaryomyces hansenii LBFT.Lev9 were able to use pectin as carbon source for their growth. On the other hand, W. cibaria LBTF.Bal3, E. thailandicus LBTF.Bal1 and W. paramesenteroides LBTF.Bal2 showed inhibitory activity against S. mutans ATCC 35668, B. subtilis subsp. spizizenii ATCC 6633 and Staph. epidermidis ATCC 14990. Results of this study are useful for the search of potential application of the isolated yeasts and lactic bacteria in coffee and other food fermentations.

Facility-specific 'house' microbiome ensures the maintenance of functional microbial communities into coffee beans fermentation: implications for source tracking.

This work aimed at studying the unconfirmed hypothesis predicting the existence of a connection between coffee farm microbiome and the resulting spontaneous fermentation process. Using Illumina-based amplicon sequencing, 360 prokaryotes and 397 eukaryotes were identified from coffee fruits and leaves, over-ripe fruits, water used for coffee de-pulping, depulped coffee beans, soil, and temporal fermentation samples at an experimental farm in Honduras. Coffee fruits and leaves were mainly associated with high incidence of Enterobacteriaceae, Pseudomonas, Colletotrichum, and Cladosporium. The proportion of Enterobacteriaceae was increased when leaves and fruits were collected on the ground compared to those from the coffee tree. Coffee farm soil showed the richest microbial diversity with marked presence of Bacillus. Following the fermentation process, microorganisms present in depulped coffee beans (Leuconostoc, Gluconobater, Pichia, Hanseniaspora, and Candida) represented more than 90% of the total microbial community, which produced lactic acid, ethanol, and several volatile compounds. The community ecology connections described in this study showed that coffee fruit provides beneficial microorganisms for the fermentation process. Enterobacteria, Colletotrichum, and other microbial groups present in leaves, fruit surface, over-ripe fruits, and soil may transfer unwanted aromas to coffee beans, so they should be avoided from having access to the fermentation tank.

Microbial fermentation affects sensorial, chemical, and microbial profile of coffee under carbonic maceration.

In view of the possibility of diversifying metabolic routes promoted by fermentation, this study proposed a new processing method for coffee, which consists of adapting a technique already consolidated in winemaking, carbonic maceration. The assay occurred under anaerobic conditions with different time and temperature fermentation. The aim of this study was to determine the differences in coffee characteristics (sensorial, chemical, and microbial) after carbonic maceration and fermentation. Specialty Coffee Association protocol, nuclear magnetic resonance, and denaturing gradient gel electrophoresis were used in these analyzes. A significant functional relationship between global score and temperature (38 °C), for the fermentation time of 96 h was observed. Bacterial diversity and sensory characteristics had a positive correlation. Furthermore, trigonelline, formic acid, hydroxymethylfurfural, lipids, and γ-butyrolactone also contributed to score and sensory quality of coffee beverage. Thus, our data show consistent factors to infer on the microbiological action on the sensory quality of coffee beverage.

The use of mesophilic and lactic acid bacteria strains as starter cultures for improvement of coffee beans wet fermentation.

The use of starter cultures during food fermentation aims to standardize the process and to obtain a higher quality product. The objectives were to study mesophilic bacteria (MB) and lactic acid bacteria (LAB) isolated from wet coffee processing and evaluate their performance in a pulped coffee medium. Eighty-six bacteria isolates (59 MB and 27 LAB) were assessed for pectinolytic activity, metabolite production, and pH value decrease in coffee-based culture (CPM). Seven bacteria strains (3 MB and 4 LAB) were selected and used as starter cultures in the wet fermentation of pulped coffee. The MB and LAB populations varied from 4.48 to 8.43 log CFU g-1 for MB and 3.54 to 8.72 log CFU g-1 for LAB during fermentation. Organic acid concentration (ranged from 0.01 to 0.53 for succinic acid; 0.71 to 8.14 for lactic acid and 0.06 to 0.29 for acetic acid), and volatile compounds (44 compounds were detected in green beans and 98 in roasted beans) were evaluated during fermentation. The most abundant compounds found in roasted beans belong to furans [15], ketones and esters [14], pyridines [13], and pyrazines [12]). Leuconostoc mesenteroides CCMA 1105 and Lactobacillus plantarum CCMA 1065 presented volatile compounds important for coffee aroma. Isovaleric acid; 2,3-butanediol; phenethyl alcohol; ß-linalool; ethyl linoleate; and ethyl 2-hydroxypropanoate could improve cupping qualities.

Levels of Alternaria Toxins in Selected Food Commodities Including Green Coffee.

Alternaria toxins are emerging mycotoxins, candidates for regulation by European Authorities. Therefore, highly sensitive, confirmatory, and reliable analytical methodologies are required for their monitoring in food. In that context, an isotope dilution LC-MS/MS method was developed for the analysis of five Alternaria toxins (Altenuene, Alternariol, Alternariol monomethylether, Tentoxin, and Tenuazonic Acid) in a broad range of commodities including cereals and cereal-based products, tomato-based products, tree nuts, vegetable oils, dried fruits, cocoa, green coffee, spices, herbs, and tea. Validation data collected in two different laboratories demonstrated the robustness of the method. Underestimation of Tenuazonic Acid level in dry samples such as cereals was reported when inappropriate extraction solvent mixtures were used as currently done in several published methodologies. An investigation survey performed on 216 food items evidenced large variations of Alternaria toxins levels, in line with data reported in the last EFSA safety assessment. The analysis of 78 green coffee samples collected from 21 producing countries demonstrated that coffee is a negligible source of exposure to Alternaria toxins. Its wide scope of application, adequate sample throughput, and high sensitivity make this method fit for purpose for the regular monitoring of Alternaria toxins in foods.

Effects of environmental factors on microbiota of fruits and soil of Coffea arabica in Brazil.

In recent years, several studies have been developed to understand the impact of fermentation on the final quality of coffee and have indicated that postharvest processing could be a determinant of quality. However, a trend has appeared as a scientific counterpoint, indicating that the interactions between soil, fruit, altitude, and slope exposures with respect to the Sun are important to understand the behavior of the microbiome in coffee. Studies on the microbiota of coffee have addressed its role during the fermentation process, however the knowledge of indigenous microorganisms harbored in fruits and soil of coffee trees growing in fields are essential, as they can contribute to fermentation. Therefore, the aim of this work was to evaluate the influence of topographic and edaphic factors on the bacterial and fungal communities present in the soil and in the fruits of Coffea arabica trees. Samples of fruits and soil were collected from different growing areas at different altitudes and soil conditions. The microbial DNA was extracted and sequenced. The results showed the contribution of environmental factors in the structure of bacterial and fungal communities. The richness, evenness and diversity of the mycobiome and bacteriome were higher in the soil than in the fruits, independent of altitude. In addition, coffee trees at higher altitudes tended to have more bacteria shared between the soil and fruits. The co-occurrence/co-exclusion network showed that bacteria-bacteria connections were greater in higher altitudes. On another hand, fungi-fungi and fungi-bacteria connections were higher in low altitudes. This was the first study that evaluates in deep the influence of environmental factors in the microbiota habiting fruits and soil coffee trees, which may affect the coffee beverage quality.

The crucial role of yeasts in the wet fermentation of coffee beans and quality.

The objective of this study was to investigate the role of yeasts in the wet fermentation of coffee beans and their contribution to coffee quality using a novel approach. Natamycin (300 ppm) was added to the fermentation mass to suppress yeast growth and their metabolic activities, and the resultant microbial ecology, bean chemistry and sensory quality were analyzed and compared to non-treated spontaneous fermentation we reported previously. The yeast community was dominated by Hanseniaspora uvarum and Pichia kudriavzevii and grew to a maximum population of about 5.5 log CFU/g in the absence of Natamycin, while when Natamycin was added yeasts were suppressed. The major bacterial species in both the spontaneous and yeast-suppressed fermentations included the lactic acid bacteria Leuconostoc mesenteroides and Lactococcus lactis, the acetic acid bacteria Gluconobacter cerinus and Acetobacter persici and the Enterobacteriaceae Enterobacter, Citrobacter and Erwinia. For both fermentations, the mucilage layers were completely degraded by the end of the process and the absence of yeast activities had no significant impact on mucilage degradation. During fermentation, reducing sugars were consumed while lactic acid was accumulated inside the beans, and its concentration was significantly higher in the spontaneous fermentation (3 times) than that where yeasts were suppressed by Natamycin. Glycerol was detected with a concentration of 0.08% in the absence of Natamycin and was not identified when Natamycin was added. Green beans fermented with yeast growth contained a higher amount of isoamyl alcohol (21 times), ethanol (3.7 times), acetaldehyde (8 times), and ethyl acetate (25 times) compared to beans fermented in the absence of yeast activities, which remained higher in the former after roasting. Beans fermented without yeast activities had a mild fruity aroma, and lower sensory scores of fragrances (7.0), flavor (6.5), acidity (6.3), body (7.0) and overall score (6.5) compared to the former. These findings demonstrated the crucial roles of yeasts in wet fermentation of coffee beans and for producing high quality coffee.

Roasted coffee wastes as a substrate for Escherichia coli to grow and produce hydrogen.

After brewing roasted coffee, spent coffee grounds (SCGs) are generated being one of the daily wastes emerging in dominant countries with high rate and big quantity. Escherichia coli BW25113 wild-type strain, mutants with defects in hydrogen (H2)-producing/oxidizing four hydrogenases (Hyd) (ΔhyaB ΔhybC, ΔhycE, ΔhyfG) and septuple mutant (ΔhyaB ΔhybC ΔhycA ΔfdoG ΔldhA ΔfrdC ΔaceE) were investigated by measuring change of external pH, bacterial growth and H2 production during the utilization of SCG hydrolysate. In wild type, H2 was produced with rate of 1.28 mL H2 (g sugar)-1 h-1 yielding 30.7 mL H2 (g sugar)-1 or 2.75 L (kg SCG)-1 during 24 h. In septuple mutant, H2 production yield was 72 mL H2 (g sugar)-1 with rate of 3 mL H2 (g sugar)-1 h-1. H2 generation was absent in hycE single mutant showing the main role of Hyd-3 in H2 production. During utilization of SCG wild type, specific growth rate was 0.72 ± 0.01 h-1 with biomass yield of 0.3 g L-1. Genetic modifications and control of external parameters during growth could lead to prolonged and enhanced microbiological H2 production by organic wastes, which will aid more efficiently global sustainable energy needs resulting in diversification of mobile and fixed energy sources.