Isolation and characterization of actinomycetes from Mural paintings of Snu- Sert-Ankh tomb, their antimicrobial activity, and their biodeterioration Microbiol. Res. (IF 2.777) Pub Date : 2018-08-15 Abeer F. Elhagrassy
A total of 35 actinomycetes were isolated from the surface of mural paintings of snu- sort- ankh in El Leasht, Egypt during four seasons all over 2016-2017. In 2009 this tomb was deteriorated by "Aspergillus niger, A. flavus, Fusarium moniliforme, Alternaria alternate, Rhizopus stolonifera, Bacillus subtilis, Bacillus cereus, and micrococcus iuteus." In 2017 the isolation of swabs presents only Aspergillus niger and about 35 actinomycetes classified to three different groups "Streptomyces, Nocardia, and Micromonospora" only five species belong to Streptomyces group showed antimicrobial activity against the previous microorganisms. These actinomycetes were identified according to their sequences in the GenBank to "Streptomyces spectabilis, S. alborgriseolus, S. globsus, S. corchorstt, S. ambofactens." In the other hand, the pigments of the wall paintings of the tomb (Egyptian blue, Egyptian green, Goethite) that analyzed by SEM-EDX, FTIR were measured by the spectrophotometer in both 2009 and 2017. The results showed that the actinomycetes could produce extracellular pigments causing a color change of archaeological pigments otherwise it helps in inhibition the growth of the previous microorganisms found in 2009. The optimization factors for increasing the antibiotic production of the Streptomyces were 3% NaCl and temperature between 30:35°c in Alkin pH (pH = 7.5).
Pseudomonas sp. AF-54 containing multiple plant beneficial traits acts as growth enhancer of Helianthus annuus L.under reduced fertilizer input Microbiol. Res. (IF 2.777) Pub Date : 2018-08-13 Afshan Majeed, M. Kaleem Abbasi, Sohail Hameed, Sumera Yasmin, Muhammad Kashif Hanif, Tahir Naqqash, Asma Imran
Plant growth promoting rhizobacteria (PGPR) are capable to increase the growth and yield of crops in eco-friendly and sustainable manner. To evaluate the response of sunflower towards inoculation with PGPR, a sunflower root associated bacterium AF-54 isolated from Diyar Gali Himalayan Mountain region, Azad Jammu and Kashmir (AJK), identified as Pseudomonas sp. by 16S rRNA sequence analysis and was characterized using polyphasic approach. The bacterium produced 23.9 µgmL-1 indole-3-acetic acid in tryptophan-supplemented medium, showed 44.28 nmoles mg-1 protein h-1 nitrogenase activity through acetylene reduction assay and released 48.80 μg mL−1 insoluble phosphorus in Pikovskaya’s broth. During P-solubilization, the pH of the Pikovskaya’s medium decreased from 7 to 3.04 due to the production of acetic acid, malic acid and gluconic acid. Pseudomonas sp. AF-54 showed metabolic versatility by utilizing79 carbon sources from BIOLOG GN2 plates and resistance to many antibiotics. Furthermore, it inhibited the growth of Fusarium oxysporum in dual culture assay. To evaluate the plant-inoculation response, series of experiments conducted in hydroponic, sterilized soil and fields at AJK, and Faisalabad where inoculated plants with reduced fertilizer showed a significant increase in growth, yield, oil contents and achene NP uptake as compared to non-inoculated control. AF-54 showed extensive root colonization in sterilized and non-sterile conditions documented through yfp-labeling and fluorescent in situ hybridization coupled with confocal laser scanning microscopy. This study concludes that the Pseudomonas sp. strain AF-54 containing multiple plant growth promoting traits can be a potential candidate for biofertilizer production to enhance sunflower crop yield with reduced application of chemical (NP) fertilizers.
Emergence of boscalid-resistant strains of Erysiphe necator in French vineyards Microbiol. Res. (IF 2.777) Pub Date : 2018-08-11 Semcheddine Cherrad, Aline Charnay, Catalina Hernandez, Herve Steva, Lassaad Belbahri, Sébastien Vacher
The grapevine powdery mildew Erysiphe necator (E. necator) is an obligate pathogen. Powdery mildew-diseased vines show an important reduction in plant size, winter hardiness and grape yield. Even a low-level infection with powdery mildew was shown to taint wine and ultimately reduce wine quality. For many years, succinate dehydrogenase inhibitor (SDHI) fungicides, mainly the new generation active ingredients (AIs) boscalid, penthiopyrad and fluopyram, have been widely used to control powdery mildew in grapevines. The repeated use of fungicides (mainly boscalid) has resulted in the emergence of resistant microorganisms such as Botrytis cinerea (B. cinerea). However, boscalid resistance was never observed in E. necator. In this study, a large-scale survey of French grapevine field populations of E. necator revealed many field populations with low sensitivity to boscalid. Single spore strains originating from collected resistant populations showed Half maximal effective concentration (EC50) values greater than 100 mg L-1, and strains originating from boscalid sensitive populations showed EC50 values lower than 1 mg L-1. The complete nucleotide sequences of the EnSdhB succinate dehydrogenase of sensitive and resistant single spore strains revealed that H242R and H242Y substitutions in the EnSdhB succinate dehydrogenase subunit conferred E. necator resistance to boscalid. No cross-resistance of E. necator strains bearing H242R and H242Y substitutions in EnSdhB succinate dehydrogenase to fluxapyroxad and fluopyram was noticed. Therefore, our results highlight the emergence of resistance to boscalid activity in French vineyards and warrant the need of the implementation of risk assessment strategies to maintain effective grapevine protection against powdery mildew.
Biological roles of indole-3-acetic acid in Acinetobacter baumannii Microbiol. Res. (IF 2.777) Pub Date : 2018-08-07 Huei-Ru Lin, Hung-Yu Shu, Guang-Huey Lin
Indole-3-acetic acid (IAA) is an important plant hormone, and many types of bacteria interact with plants by producing or degrading IAA in the rhizosphere. The iac (indole-3-acetic acid catabolism) gene locus in Acinetobacter baumannii ATCC19606 was previously associated with IAA degradative capability, and in this study, transcriptome analysis results derived from A. baumannii cultured with IAA showed that the expression of catechol-degrading and phenylacetate-degrading genes was elevated, indicating that IAA is likely degraded through these pathways. This study further found that A. baumannii also has IAA productive capability, primarily involving the ipdC gene, and transcriptome and spent media analysis of wild-type and mutant cultures grown in minimal media revealed that A. baumannii likely produces IAA through the indole-3-pyruvic acid (IPyA) pathway. Exogenously applied IAA improved tolerance against oxidative stress in wild-type A. baumannii and iacA mutants unable to degrade IAA, but not in ipdC mutants incapable of producing IAA, suggesting that endogenous IAA is important for stress tolerance. Meanwhile, ipdC mutants also had reduced virulence against human A549 epithelial cells as compared to wild-type. Endogenously produced IAA was found to enhance root growth in A. baumannii and kidney bean plant co-cultures, indicating that A. baumannii can interact with plants through the production and degradation of IAA. Taken together, this study sheds light on the biosynthesis pathways and functional significance of IAA in A. baumannii, and may be useful in exploring other IAA-mediated plant-microbe interactions as well.
Phenotype responses to abiotic stresses, asexual reproduction and virulence among isolates of the entomopathogenic fungus Cordyceps javanica (Hypocreales: Cordycipitaceae) Microbiol. Res. (IF 2.777) Pub Date : 2018-08-04 Gabriel Moura Mascarin, Ronaldo Alves Pereira-Junior, Éverton Kort Kamp Fernandes, Eliane Dias Quintela, Christopher A. Dunlap, Steven Paul Arthurs
Growth and protease secretion of Scedosporium aurantiacum under conditions of hypoxia Microbiol. Res. (IF 2.777) Pub Date : 2018-08-04 Zhiping Han, Liisa Kautto, Wieland Meyer, Sharon C.-A. Chen, Helena Nevalainen
One of the micro-environmental stresses that fungal pathogens, such as Scedosporium aurantiacum, colonising human lungs encounter in vivo is hypoxia, or deficiency of oxygen. In this work, we studied the impacts of a hypoxic micro-environment (oxygen levels ≤1%) on the growth of a clinical S. aurantiacum isolate (WM 06.482; CBS 136046) and an environmental strain (S. aurantiacum WM 10.136; CBS 136049) on mucin-containing synthetic cystic fibrosis sputum medium. Additionally, profiles of secreted proteases were compared between the two isolates and protease activity was assessed using class-specific substrates and inhibitors. Overall, both isolates grew slower and produced less biomass under hypoxia compared to normoxic conditions. The pH of the medium decreased to 4.0 over the cultivation time, indicating that S. aurantiacum released acidic compounds into the medium. Accordingly, secreted proteases of the two isolates were dominated by acidic proteases, including aspartic and cysteine proteases, with optimal protease activity at pH 4.0 and 6.0 respectively. The clinical isolate produced higher aspartic and cysteine protease activities. Conversely, all serine proteases, including elastase-like, trypsin-like, chymotrypsin-like and subtilisin-like proteases had higher activities in the environmental isolate. Sequence similarities to 13 secreted proteases were identified by mass spectrometry (MS) by searching against other fungal proteases in the NCBI database. Results from MS analysis were consistent with those from activity assays. The clinical highly-virulent, and environmental low-virulence S. aurantiacum isolates responded differently to hypoxia in terms of the type of proteases secreted, which may reflect their different virulence properties.
The first report of antifungal lipopeptide production by a Bacillus subtilis subsp. inaquosorum strain Microbiol. Res. (IF 2.777) Pub Date : 2018-08-02 Chinyere Knight, Michael J. Bowman, Lafayette Frederick, Agnes Day, Clarence Lee, Christopher A. Dunlap
A strain of bacteria in the Bacillus subtilis species complex was isolated from a building’s air vent in the Washington DC area, USA, and produced strong antifungal activity with in vitro assays. This strain, designated (HU Biol-II), showed pronounced inhibitory effects on mycelial growth of a wide spectrum of fungi. The objectives of this study were to use genome sequencing to confirm the taxonomy of HU Biol-II, evaluate its antifungal activity and implement genome mining and HPLC-MS/MS to characterize the bioactive secondary metabolites. The strain, as determined by multilocus sequence alignment analysis, was identified as a member of Bacillus subtilis subsp. inaquosorum clade. Core genome phylogeny showed that the isolate is most closely related to B. subtilis subsp. inaquosorum strain DE111, a commercially produced human probiotic. The investigation identified eight bioactive metabolite clusters in the genome. HPLC MS/MS was able to confirm the production of seven of the metabolites. This study is the first to report the production of two antifungal cyclic lipopeptides (bacillomycin F and fengycin) from a member of B. subtilis subsp. inaquosorum. The strain also produced the antibacterial aurantinin B, which confirms the biosynthetic cluster responsible for its production. Comparative genomics and metabolomics demonstrated the commercial probiotic strain DE111 produced the same metabolites, with the exception of aurantinin B. These findings are the first description of the secondary metabolites produced by a strain of B. subtilis subsp. inaquosorum.
Insights into the carbonic anhydrases and autotrophic carbon dioxide fixation pathways of high CO2 tolerant Rhodovulum viride JA756 Microbiol. Res. (IF 2.777) Pub Date : 2018-07-20 Lakshmi Venkata Naga Satya Khandavalli, Tushar Lodha, Maaged Abdullah, Lalitha Guruprasad, Sasikala Chintalapati, Venkata Ramana Chintalapati
Biofixation of CO2 is being extensively investigated to solve the global warming problem. Purple non-sulfur bacteria are fast growers that consume CO2 and produce beneficial biomass. Better the growth at higher CO2 levels, more efficient are the strains for biofixation. Nine among fifty strains that were analyzed at elevated CO2 levels responded with better growth. Considering its enhanced growth at high CO2 and metabolic versatility, Rhodovulum viride strain JA756 was chosen to make further studies. Strain JA756 tolerates up to 50% (v/v) CO2 with its optimum between 20–40% (v/v), yielding a biomass of 3.4 g. L−1. The pattern of specific enzyme activity of carbonic anhydrase corresponded well with that of its growth. To gain insights into the genomic composition and genes related to carbonic anhydrases and CO2 fixation, draft genome sequencing of JA756 was carried out which revealed the presence of two non-homologous genes encoding for β and γ carbonic anhydrases, both of which are assumed to be implicated in maintaining intracellular inorganic carbon concentration at equilibrium. Most of the genes involved in the Calvin pathway, reductive tricarboxylic acid pathway, 3-hydroxypropionate bicycle and C4 pathways were found in the draft genome. While the experimental determinations of active roles of two of these pathways are still underway, the expression of key genes of Calvin and C4 pathway suggest their functional role in the organism. Owing to its metabolic versatility, JA756 can be advantageous for biological CO2 assimilation facilities located by the coastline, inland and also at wide ranges of CO2 concentrations.
Soil amendment alters soil physicochemical properties and bacterial community structure of a replanted apple orchard Microbiol. Res. (IF 2.777) Pub Date : 2018-07-20 Bowen Liang, Changqing Ma, Lianmei Fan, Yongzhang Wang, Yongbing Yuan
Compost amendment reportedly improved apple tree growth in replant soils. However, its effects should be evaluated at different soil depths and locations. This study investigated the impact of soil improvement with compost on soil physicochemical properties and bacterial community structure of a replanted apple orchard in comparison with the original orchard without compost improvement. The V1–V3 region of the bacterial 16S rRNA gene was subjected to high-throughput 454 pyrosequencing, and data were analyzed using the Mothur pipeline. The results showed that the soil improvement benefited tree growth and fruit quality during the study period. The compost amendment markedly increased tree height and stem diameter by a range of 6.1% to 21.0% and 4.0% to 14.0%, respectively. Fruit yield (9.5%), average weight (9.6%), and soluble solid content (5.6%) were also increased by compost amendment compared to those of the unimproved treatment. The pH, organic matter, and available N, P, and K contents were significantly increased by 5.7% to 21.9%, 0.2% to 62.9%, 9.3% to 29.3%, 36.7% to 64.5%, and 17.2% to 100.3% in the compost improved soil. The pyrosequencing data showed that the soil improvement changed the bacterial community structure at all soil depths (0–20 cm and 20–40 cm) and locations (in-row and inter-row) considered; e.g., the relative abundance of Proteobacteria (20.2%), Bacteroidetes (2.5%), and Cyanobacteria (1.0%) was increased while that of Chloroflexi (5.5%), Acidobacteria (5.2%), Nitrospirae (4.5%), Gemmatimonadetes (3.8%), and Actinobacteria (1.8%) was decreased. The relative abundance of some dominant genera Burkholderia (2.3%), Pseudomonas (1.0%), and Paenibacillus (0.5%) were enhanced in the compost improved soil. Moreover, other dominant genera such as Nitrospira (6.4%), Gemmatimonas (2.2%), and Phenylobacterium (0.3%) were reduced by the application of compost. Our results indicate that soil improvement benefits the growth of tree and fruit quality, and is likely mediated by increased soil pH, organic matter, and available nutrient contents and beneficial bacterial community composition.
The role of two-component regulatory system in β-lactam antibiotics resistance Microbiol. Res. (IF 2.777) Pub Date : 2018-07-19 Li Lingzhi, Ge Haojie, Gu Dan, Meng Hongmei, Li Yang, Jia Mengdie, Zheng Chengkun, Zhou Xiaohui
The irrational use of antibiotics in agriculture and in the medical field has led to a variety of pathogenic microorganisms that produce drug resistance and even multidrug resistance. B-lactam is one of the most widely used antibiotics to treat infectious diseases. Resistance to β-lactam resistance can be primarily due to the presence β-lactamase, mutation of β-lactam targets and overexpression of efflux pumps. Two-component regulatory systems are composed of histidine kinase and response regulator that regulate gene expression under different environmental conditions. In this review, we summarized the mechanisms by which β-lactam resistance is developed and the role of the two-component regulatory system in β-lactam resistance.
Paenibacillus lentimorbus induces autophagy for protecting tomato from Sclerotium rolfsii infection Microbiol. Res. (IF 2.777) Pub Date : 2018-07-19 Ritu Dixit, Lalit Agrawal, Surendra Pratap Singh, Prateeksha, Poonam C. Singh, Vivek Prasad, Puneet Singh Chauhan
During biotic stress, plants use several mechanisms to protect themselves that include the production of reactive oxygen species (ROS), induction of pathogenesis-related proteins and cell death. Some plant growth promoting rhizobacteria (PGPR) are known to act as bio-control agents that protect crops against pathogens. The biocontrol activity of PGPR Paenibacillus lentimorbus (B-30488) against Sclerotium rolfsii showed previously where several defense-related genes were upregulated with ROS induction in tomato. We further evaluate the other possibility, i.e. role of autophagy in enhancing defense in tomato using PGPR. Confocal microscopy revealed the presence of an acidotropic dye Mono Dansyl Cadaverine (MDC) stained autophagosomes in B-30488 treated healthy and infected plants. These autophagosomes almost disappeared in plants treated with an autophagy inhibitor chloroquine. The results were also confirmed by ultrastructural analysis of leaf tissues using transmission electron microscopy. Enhanced expression of autophagy-related genes was also monitored in B-30488 primed fungal infected tissues as compared to control by qRT-PCR. Results of ROS accumulation, fluorescence, confocal and transmission electron microscopy and gene expression analysis revealed induction of autophagy using B-30488 as a biocontrol agent suggesting a role in enhancing disease resistance in tomato. Overall, the present study indicated a role of B-30488 as a biocontrol in enhancing disease resistance in tomato and also assists a better understanding of fungal pathogenesis that is expected to be useful in developing new strategies for disease control.
Versatile use of Azospirillum brasilense strains tagged with egfp and mCherry genes for the visualization of biofilms associated with wheat roots Microbiol. Res. (IF 2.777) Pub Date : 2018-07-19 Alberto Ramirez-Mata, Miguel Ramales Pacheco, Saul Jijon Moreno, Maria Luisa Xiqui-Vazquez, Beatriz E. Baca
This study reports the introduction of egfp or mCherry markers to the Sp245, Sp7, and M40 wild-type strains of Azospirillum brasilense and the hhkB (encoding for a putative hybrid histidine kinase) minus mutant an isogenic strain of A. brasilense Sp245 to monitor colonization of wheat (Triticum aestivum). Two plasmids were constructed: (1) the pJMS-2 suicide plasmid derived from pSUP202 and harboring the mCherry gene expressed under the constitutive kanamycin resistance promoter to create a cis tag and (2) the broad-range plasmid pMP2449-5 that carries the mCherry gene under the lac promoter, which is derived from the plasmid pMP2444; to create the in trans tag. The stability of the plasmids encoding egfp and mCherry were confirmed in vitro for seven days of bacterial growth, and then, the A. brasilense strains harboring the plasmids were studied under nonselective conditions for adherence to seeds and, at seven or 14 days post-inoculation, for wheat root colonization. The utility of the labeled strains was proven by observation, using fluorescence microscopy and confocal laser scanning microscopy (CLSM) in wheat plants inoculated with the labeled strains and compared with the CFU g–1 for seed and wheat root. The method was suitable for observation of the in situ formation of mini-colonies, enabled visualization of bacterial colonization sites on large root fragments, and showed adherence to germinated seeds and root colonization of all strains by cell counts and direct microscopic examination. Thus, we are able to quantify the structures of the biofilms formed by each strain.
A novel flow-system to establish experimental biofilms for modelling chronic wound infection and testing the efficacy of wound dressings Microbiol. Res. (IF 2.777) Pub Date : 2018-07-19 Peter F. Duckworth, Richard S. Rowlands, Michele E. Barbour, Sarah E. Maddocks
Several models exist for the study of chronic wound infection, but few combine all of the necessary elements to allow high throughput, reproducible biofilm culture with the possibility of applying topical antimicrobial treatments. Furthermore, few take into account the appropriate means of providing nutrients combined with biofilm growth at the air-liquid interface. In this manuscript, a new biofilm flow device for study of wound biofilms is reported. The device is 3D printed, straightforward to operate, and can be used to investigate single and mixed species biofilms, as well as the efficacy of antimicrobial dressings. Single species biofilms of Staphylococcus aureus or Pseudomonas aeruginosa were reproducibly cultured over 72 h giving consistent log counts of 8-10 colony forming units (CFU). There was a 3-4 log reduction in recoverable bacteria when antimicrobial dressings were applied to biofilms cultured for 48 h, and left in situ for a further 24 h. Two-species biofilms of S. aureus and P. aeruginosa inoculated at a 1:1 ratio, were also reproducibly cultured at both 20 °C and 37 °C; of particular note was a definitive Gram-negative shift within the population that occurred only at 37 °C.
Specific growth inhibitors of Ralstonia solanacearum, Xanthomonas oryzae pv. oryzae, X. campestris pv. campestris, and Clavibacter michiganensis subsp. michiganensis Microbiol. Res. (IF 2.777) Pub Date : 2018-06-10 Geofrey Sing’ombe Ombiro, Taku Sawai, Yoshiteru Noutoshi, Yuta Nishina, Hidenori Matsui, Mikihiro Yamamoto, Kazuhiro Toyoda, Yuki Ichinose
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-β-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.
Bacteria and bacterial products: Foe and friends to Caenorhabditis elegans Microbiol. Res. (IF 2.777) Pub Date : 2018-06-25 Fazlurrahman Khan, Saurabh Jain, Sandra Folarin Oloketuyi
Caenorhabditis elegans is a model organism for the study of different molecular, biochemical, microbial and immunity-related mechanisms. In its natural habitat, C. elegans survives by feeding microorganisms (mainly bacteria), though majorly on Escherichia coli OP50 when grown in the laboratory. Numerous bacteria are shown to influence the lifespan, behavioural responses and innate immunity of C. elegans. The secondary metabolites produced by bacteria have shown to play key role in C. elegans longevity. This behaviour provides insights for potential development of new strategies for the treatment of diseases in other species, including humans. This review explains the concept of C. elegans microbiome, different mechanisms employed in its longevity and resistance against bacterial pathogens and the effects of various bacteria (both beneficial and harmful) as well as their products on the life cycle of C. elegans.
Designer probiotic Lactobacillus plantarum expressing oxalate decarboxylase developed using group II intron degrades intestinal oxalate in hyperoxaluric rats Microbiol. Res. (IF 2.777) Pub Date : 2018-06-22 Eldho Paul, Abhishek Albert, Sasikumar Ponnusamy, Srishti Rajkumar Mishra, Amalraj Ganesh Vignesh, Selvi Mariaraj Sivakumar, Gomathi Sivasamy, Selvam Govindan Sadasivam
Increased intestinal absorption of oxalate causes hyperoxaluria, a major risk factor for kidney stone disease. Intestinal colonization of recombinant probiotic bacteria expressing oxalate-degrading gene (OxdC) is an effective therapeutic option for recurrent calcium oxalate (CaOx) stone disease. Therefore, we aimed to develop food-grade probiotic L. plantarum secreting OxdC using lactococcal group II intron, Ll.LtrB and evaluate its oxalate degradation ability in vivo. Male Wistar albino rats were divided into four groups. The rats of group I received normal rat chow and drinking water. Groups II, III and IV rats received 5% potassium oxalate containing diet for 28 days. Groups III and IV rats received L. plantarum and food-grade recombinant L. plantarum respectively from 15 to 28 days. Biochemical parameters and crystalluria were analysed in 24 h urine samples. At the end of experimental period, rats were sacrificed; intestine and kidneys were dissected out for colonization studies and histopathological analysis. Herein, we found that the administration of recombinant probiotics significantly reduced the urinary oxalate, calcium, urea, and creatinine levels in rats of group IV compared to group II. Furthermore, colonization studies indicated that recombinant probiotics have gastrointestinal transit and intestinal colonization ability similar to that of wild-type bacteria. In addition, gene expression studies revealed down-regulation of OPN and KIM-1 among group IV rats. Histopathological analysis showed less evidence of nephrocalcinosis in group IV rats. In conclusion, the study demonstrates that food-grade L. plantarum secreting OxdC is capable of degrading intestinal oxalate and thereby prevent CaOx stone formation in experimental rats.
Pyrrolnitrin is more essential than phenazines for Pseudomonas chlororaphis G05 in its suppression of Fusarium graminearum Microbiol. Res. (IF 2.777) Pub Date : 2018-06-18 Run Huang, Zhibin Feng, Xiaoyan Chi, Xiaoqiang Sun, Yang Lu, Baoshen Zhang, Ruiyang Lu, Wangtai Luo, Yanhua Wang, Jing Miao, Yihe Ge
Fusarium graminearum is the major causal agent of Fusarium head blight (FHB) disease in cereal crops worldwide. Infection with this fungal phytopathogen can regularly cause severe yield and quality losses and mycotoxin contamination in grains. In previous other studies, one research group reported that pyrrolnitrin had an ability to suppress of mycelial growth of F. graminearum. Other groups revealed that phenazine-1-carboxamide, a derivative of phenazine-1-carboxylic acid, could also inhibit the growth of F. graminearum and showed great potentials in the bioprotection of crops from FHB disease. In our recent work with Pseudomonas chlororaphis strain G05, however, we found that although the phz operon (phenazine biosynthetic gene cluster) was knocked out, the phenazine-deficient mutant G05Δphz still exhibited effective inhibition of the mycelial growth of some fungal phytopathogens in pathogen inhibition assay, especially including F. graminearum, Colletotrichum gloeosporioides, Botrytis cinerea. With our further investigations, including deletion and complementation of the prn operon (pyrrolnitrin biosynthetic gene cluster), purification and identification of fungal compounds, we first verified that not phenazines but pyrrolnitrin biosynthesized in P. chlororaphis G05 plays an essential role in growth suppression of F. graminearum and the bioprotection of cereal crops against FHB disease.
Secretome analysis of alkaliphilic bacterium Bacillus lehensis G1 in response to pH changes Microbiol. Res. (IF 2.777) Pub Date : 2018-06-19 How Lie Ling, Zaidah Rahmat, Farah Diba Abu Bakar, Abdul Munir Abdul Murad, Rosli Md. Illias
Bacillus lehensis G1 is an alkaliphilic bacterium that is capable of surviving in environments up to pH 11. Secretome related to bacterial acclimation in alkaline environment has been less studied compared to cytoplasmic and membrane proteome. The aim of this study was to gain better understanding of bacterial acclimation to alkaline media through analyzing extracellular proteins of B. lehensis. The pH range for B. lehensis growth was conducted, and two-dimensional electrophoresis and MALDI-TOF/TOF MS analysis were conducted to characterize changes in protein profiling in B. lehensis cultured at pH 8 and pH 11 when compared with those cultured at pH 10 (optimal growth pH). B. lehensis could grow well at pH ranging from 8 to 11 in which the bacteria showed to posses thinner flagella at pH 11. Proteomic analyses demonstrated that five proteins were up-regulated and 13 proteins were down-regulated at pH 8, whereas at pH 11, 14 proteins were up-regulated and 8 were down-regulated. Majority of the differentially expressed proteins were involved in the cell wall, main glycolytic pathways, the metabolism of amino acids and related molecules and some proteins of unknown function. A total of 40 differentially expressed protein spots corresponding to 33 proteins were identified; including GlcNAc-binding protein A, chitinase, endopeptidase lytE, flagellar hook-associated proteins and enolase. These proteins may play important roles in acclimation to alkaline media via reallocation of cell wall structure and changes to cell surface glycolytic enzymes, amino acid metabolism, flagellar hook-associated proteins and chaperones to sustain life under pH-stressed conditions.
Antibiotic producing endophytic Streptomyces spp. colonize above-ground plant parts and promote shoot growth in multiple healthy and pathogen-challenged cereal crops Microbiol. Res. (IF 2.777) Pub Date : 2018-06-03 Janki K. Patel, Sheeba Madaan, G. Archana
The Streptomyces spp. used in this work were previously isolated as diazotrophic endophytes from sorghum stems. Here, we characterized the Streptomyces spp. for their colonization ability, plant growth promotion and protection against fungal disease in three cereals. In vitro analysis by dual culture study showed inhibitory effect on the rice pathogen Magnaporthe oryzae B157 along with inhibition of the ubiquitous phytopathogen Rhizoctonia solani by the Streptomyces spp. used in this study. The active compounds responsible for phytopathogen inhibition were extracted with ethyl acetate and tested positive against the fungal pathogens. GC–MS based identification of the active compounds responsible for fungal pathogen inhibition showed them to be 2-(chloromethyl)-2-cyclopropyloxirane, 2, 4- ditert-butylphenol and 1‐ethylthio‐3‐methyl‐1, 3‐butadiene in extracts of culture supernatants from the three different strains respectively. EGFP tagged Streptomyces strains showed profuse colonization in roots as well as aerial parts of cereal plants. Direct inhibitory action against M. oryzae B157 and R. solani correlated with the observation that upon fungal pathogen challenge, the bacterized rice, sorghum and wheat plants showed significantly good plant growth, particularly in aerial parts as compared to unbacterized controls. In addition, benefit was seen in inoculated healthy plants in terms of increase in wet weight of roots and shoots as compared to the uninoculated controls. The mechanism of biocontrol also involved induction of plant defense response as evidenced by the upregulation of PR10a, NPR1, PAL and LOX2 in Streptomyces colonized plants.
Matricaria genus as a source of antimicrobial agents: From farm to pharmacy and food applications Microbiol. Res. (IF 2.777) Pub Date : 2018-06-25 Mehdi Sharifi-Rad, Jolanta Nazaruk, Letizia Polito, Maria Flaviana Bezerra Morais-Braga, Janaína Esmeraldo Rocha, Henrique Douglas Melo Coutinho, Bahare Salehi, Giulia Tabanelli, Chiara Montanari, María del Mar Contreras, Zubaida Yousaf, William N. Setzer, Deepa R. Verma, Miquel Martorell, Antoni Sureda, Javad Sharifi-Rad
Matricaria is a widespread genus of flowering plants of the family Asteraceae that grow in temperate regions of Europe, Asia, America and Africa. Some of the species are also naturalized in Australia. Some species of this genus such as Chamomiles are recognized medicinal plants and cultivated in several countries for commercial purposes: to obtain its blue essence, as herbal tea, and for pharmaceutical or cosmeceutical uses. The phytochemical composition of Matricaria spp. includes volatile terpenoids (e.g., α-bisabolol, bisabolol oxide A and B, β-trans-farnesene and chamazulene), sesquiterpene lactones such as matricin, and phenolic compounds (flavonoids, coumarins and phenolic acids). Their essential oil is obtained from the fresh or dried inflorescences by steam distillation, and additionally cohobation of the remaining water. The volatile composition of the essential oil, especially the content of the valuable components α-bisabolol and chamazulene, depends on the plant part, origin and quality of the source, genetic, and environmental factors. Moreover, other parameters, such as season of harvest and methods of extraction, can affect the extraction yield of the essential oils/extracts, their composition and, therefore, their bioactivity. Due to the importance of this genus and particularly M. recutita (M. chamomilla), this review focus on its cultivation, factor affecting essential oils’ composition and their role in traditional medicine, as antibacterial agents and finally as food preservatives.
Bacillus thuringiensis produces the lipopeptide thumolycin to antagonize microbes and nematodes Microbiol. Res. (IF 2.777) Pub Date : 2018-06-03 Dehong Zheng, Zhiyong Zeng, Bingbing Xue, Yaoyao Deng, Ming Sun, Ya-Jie Tang, Lifang Ruan
Bacillus thuringiensis has been widely used as a bio-insecticide. However, novel biological activities other than insect toxicity of B. thuringiensis are still underestimated. In this study, a new lipopeptide biosynthesis gene cluster in B. thuringiensis BMB171 was discovered by genome mining and verified by reverse genetics. Thumolycin, the lipopeptide synthesized by this gene cluster, was then isolated and purified. Mass spectrum analysis revealed the molecular mass of thumolycin is 696.51 Da with the predicted molecular formula of C38H64N8O4. Further bioactivities assay showed that thumolycin endowed B. thuringiensis BMB171 with broad spectrum antimicrobial and nematocidal activities.
Hydroxycoumarins: New, effective plant-derived compounds reduce Ralstonia pseudosolanacearum populations and control tobacco bacterial wilt Microbiol. Res. (IF 2.777) Pub Date : 2018-05-17 Liang Yang, Lintong Wu, Xiaoyuan Yao, Shiyuan Zhao, Jiao Wang, Shili Li, Wei Ding
Plant wilt disease caused by the soilborne bacterial pathogen Ralstonia pseudosolanacearum is one of the most devastating plant diseases; however, no effective protection against this disease has been developed. Coumarins are important natural plant-derived compounds with a wide range of bioactivities and extensive applications in medicine and agriculture. In the present study, three hydroxycoumarins (Hycs), umbelliferone (UM), esculetin (ES) and daphnetin (DA) significantly inhibited the growth of R. pseudosolanacearum on solid medium in a concentration-dependent manner, and the minimum inhibitory concentration (MICs) of these compounds was 325 mg L−1, 125 mg L−1 and 75 mg L−1, respectively. The percentage of live cells of R. pseudosolanacearum when supplemented with UM, ES, and DA was 63.61%, 17.81% and 7.23%, respectively, which were significantly lower than the DMSO treatment with 92%. Furthermore, irrigating roots with hydroxycoumarins (Hycs) 24 h before inoculation with R. pseudosolanacearum significantly delayed the occurrence of tobacco bacterial wilt, with the control efficiency of the DA treatment (the most efficient of Hycs treatment) 80.03%, 69.83%, 59.19%, 45.49%, 44.12%, 38.27% at 6, 8, 10, 12, 14, and 16 days after inoculation, respectively. Compared with the DMSO treatment, the pathogen populations of tobacco stems supplemented with 100 mg L−1 DA were the lowest, with population significantly reduced by 22.46%, 27.34%, and 18.06% at 4, 7, and 10 days after inoculation, respectively. Based on this study, these Hycs could be applied as potential protective agents in the management of tobacco bacterial wilt.
Current insight and futuristic vistas of microbial transglutaminase in nutraceutical industry Microbiol. Res. (IF 2.777) Pub Date : 2018-06-04 Syeda Warisul Fatima, Sunil K. Khare
Microbial transglutaminase (MTGase) has become a driving force in the food industry cross-linking the food proteins. MTGase-the nature’s molecular glue is recognized to reorient food protein’s functional properties without affecting its nutritive value. The scope and approach of this review is to have insight on the action mechanism of MTGase and impact of molecular linkage on functional proteins in various protein moieties in development of innovative features in food production for better consumer’s choice and satisfaction. The study covers a wide range of published work across food industries involving innovative use of MTGase, an environment friendly production approach for commercial utilization to get better outcome in terms of culinary delight. The intrinsic biochemical properties and structural information by sequence analysis and clustering validates the mode of reaction mechanism of the biological glue enzyme. The review singles out how the MTGase emerged as a prime choice in ever evolving food industry.
Inhibition of biofilm formation by Cd2+ on Bacillus subtilis 1JN2 depressed its biocontrol efficiency against Ralstonia wilt on tomato Microbiol. Res. (IF 2.777) Pub Date : 2018-06-02 Wei Yang, Haixia Yan, Ji Zhang, Yuming Gao, Wei Xu, Jinshan Shang, Yuming Luo
Bacillus subtilis 1JN2 can serve as an effective biocontrol agent against Ralstonia wilt on tomato, but the efficiency of control depends on the levels of heavy metals in the rhizosphere soil. Here, we investigated how the heavy metal Cd2+ affects the biocontrol efficacy of B.subtilis 1JN2 on Ralstonia wilt. We found that low Cd2+ content of 2 mM or lower had no effects on the biofilm formation of 1JN2, while media containing 3 mM or higher Cd2+ levels inhibited biofilm formation. Interestingly, high concentration of Cd2+ (5 mM) showed inhibition of B.subtilis 1JN2 cell growth. We next tested the effects of Cd2+ on the colonization of 1JN2 by supplementing artificial Cd2+ in the tomato rhizosphere in a greenhouse setting. We found that 3 mM Cd2+ in the tomato rhizosphere inhibited the colonization of B.subtilis 1JN2, Only 103 CFU/mL 1JN2 was detected one week post treated with 107 CFU/mL but 105 CFU/mL could be detected without Cd2+ in the soil. The presence of Cd2+ had no effect on the colonization of Ralstonia solanacearum on tomato, but the biocontrol efficacy against Ralstonia wilt by 1JN2 decreased 54.2% when the soil contained 3 mM Cd2+ compared to the control without Cd2+. Taken together, we found that the failure of biofilm formation of Bacillus subtilis 1JN2 that affected by Cd2+ lead to the decrease of its biocontrol efficacy against Ralstonia wilt on tomato.
Brevibacterium linens RS16 confers salt tolerance to Oryza sativa genotypes by regulating antioxidant defense and H+ ATPase activity Microbiol. Res. (IF 2.777) Pub Date : 2018-06-19 Poulami Chatterjee, Sandipan Samaddar, Ülo Niinemets, Tong-Min Sa
Soil salinity is one of the major limitations that affects both plant and its soil environment, leading to reduced agricultural production. Evaluation of stress severity by plant physical and biochemical characteristics is an established way to study plant-salt stress interaction, but the halotolerant properties of plant growth promoting bacteria (PGPB) along with plant growth promotion is less studied till date. The aim of the present study was to elucidate the strategy, used by ACC deaminase-containing halotolerant Brevibacterium linens RS16 to confer salt stress tolerance in moderately salt-tolerant (FL478) and salt-sensitive (IR29) rice (Oryza sativa L.) cultivars. The plants were exposed to salt stress using 0, 50, and 100 mM of NaCl with and without bacteria. Plant physiological and biochemical characteristics were estimated after 1, 5, 10 days of stress application. H+ ATPase activity and the presence of hydroxyectoine gene (ectD) that is responsible for compatible solute accumulation were also analyzed in bacteria. The height and dry mass of bacteria inoculated plants significantly increased compared to salt-stressed plants, and the differences increased in time dependent manner. Bacteria priming reduced the plant antioxidant enzyme activity, lipid peroxidation and it also regulated the salt accumulation by modulating vacuolar H+ ATPase activity. ATPase activity and presence of hydroxyectoine gene in RS16 might have played a vital role in providing salt tolerance in bacteria inoculated rice cultivars. We conclude that dual benefits provided by the halotolerant plant growth promoting bacteria (PGPB) can provide a major way to improve rice yields in saline soil.
Comparison of volatile compounds released by entomopathogenic fungi Microbiol. Res. (IF 2.777) Pub Date : 2018-06-24 Aleksandra Bojke, Cezary Tkaczuk, Piotr Stepnowski, Marek Gołębiowski
Entomopathogenic fungi are fungal species which are used as a potential source of biopesticides. These fungi produce secondary metabolites which in insects can cause disruption in the normal functioning of their bodies, disease or even death. In order to fully characterize the physiology of entomopathogenic fungi we should identify the volatile organic compounds which are involved in this process. Therefore, we conducted a qualitative and quantitative analysis of volatile compounds produced by entomopathogenic fungi. Seven different species of fungi were analyzed: Metarhizium anisopliae, Metarhizium flavoviride, Pandora sp., Isaria fumosorosea, Hirsutella danubiensis, Batkoa sp. and Beauveria bassiana. The analyses were performed using the HS-SPME/GCMS technique. In the analyzed fungi, 63 volatile compounds were identified and classified into the following groups: aldehydes, ketones, alcohols, esters, acids, terpenes and others. The results show that entomopathogenic fungi produce a wide profile of secondary metabolites. Principal Components Analysis was used to determine whether separate classes of fungi can be distinguished from one another based on their metabolite profiles.
Role of two-component regulatory systems in the virulence of Streptococcus suis Microbiol. Res. (IF 2.777) Pub Date : 2018-07-07 Chengkun Zheng, Lingzhi Li, Haojie Ge, Hongmei Meng, Yang Li, Weicheng Bei, Xiaohui Zhou
Streptococcus suis is an important zoonotic pathogen that causes severe infections and great economic losses worldwide. Understanding how this pathogen senses and responds to environmental signals during the infectious process can offer insight into its pathogenesis and may be helpful in the development of drug targets. Two-component regulatory systems (TCSs) play an essential role in this environmental response. In S. suis, at least 15 groups of TCSs have been predicted. Among them, several have been demonstrated to be involved in virulence and/or stress response. In this review, we discuss the progress in the study of TCSs in S. suis, focusing on the role of these systems in the virulence of this bacterium.
A toxin-antitoxin system is essential for the stability of mosquitocidal plasmid pBsph of Lysinibacillus sphaericus Microbiol. Res. (IF 2.777) Pub Date : 2018-06-28 Pan Fu, Yong Ge, Yimin Hu, Zhiming Yuan, Xiaomin Hu
Lysinibacillus sphaericus C3-41 carries a large low-copy-number plasmid pBsph, which encodes binary toxin proteins. Our previous study found that the transcriptional activator TubX plays an important role in the newly identified type Ⅲ TubRZC replication/partition system in pBsph, and that a vector consisting of tubRZC and tubX is not as stable as pBsph, indicating the presence of other maintenance module(s). In this study, we identified that orf9 and orf10 are necessary for the stability of pBsph by a series of deletion and complementation experiments. Bioinformatics analysis showed that ORF9 contains a PIN domain of VapBC toxin-antitoxin (TA) system, whereas ORF10 share no significant sequence similarity to any of the characterized antitoxins in the database. Further studies revealed that orf9 and orf10 are transcribed as an operon. The overexpression of ORF9 repressed the growth of both Escherichia coli and L. sphaericus, which can be alleviated by overexpression of ORF10. The deletion of orf10 individually or orf9-10 together resulted a decrease on plasmid stability which was restored by the complementation of corresponding gene(s), suggesting that ORF10 plays an important role in plasmid stability. In addition, it was found the plasmid stability is related with the transcription level of tubRZ, and overexpression of TubRZ could neutralize the negative effect on plasmid stability caused by the deletion of orf9-orf10. Moreover, the recombinant vector containing tubRZC, tubX and orf9-10 was more stable than the ones containing only tubRZC and either tubX or orf9-10. The data indicate that the plasmid maintenance system on pBsph includes orf9-orf10 TA system.
Growth stage and tissue specific colonization of endophytic bacteria having plant growth promoting traits in hybrid and composite maize (Zea mays L.) Microbiol. Res. (IF 2.777) Pub Date : 2018-05-26 Premsing Shivsing Marag, Archna Suman
Maize, a crop cultivated worldwide, was investigated for plant tissue and crop stage specific colonization of endophytic bacteria. Such bacterial interactions have high potential to enhance maize grain yield by means of biological nitrogen fixation and/or plant growth promoting activities. In this study endophytic bacteria were isolated from a hybrid PEEHM-5 and composite PC-4 maize varieties using root, stem and leaf tissues of plants at vegetative, flowering and maturity stages of growth. PEEHM-5 harbored higher endophytic bacterial population than PC-4 at all growth stages, with highest in roots and at flowering stage. Morphologically 188 different endophytic isolates (82 from PEEHM-5, 106 from PC-4) were screened for plant growth promoting attributes viz. P, K, Zn solubilization, production of hormones, siderophore, ACC deaminase, HCN, biological nitrogen fixation and biocontrol of two maize fungal pathogens. Thirty one potential PGP isolates on RFLP analysis of their amplified 16S rRNA gene, were clustered in 13 phylogenetic groups. On sequencing and blasting of amplified 16S rRNA gene of representative isolates from each group identified PC-4 endophytic bacterial isolates as Bacillus aryabhattai, Pantoea cypripedii, Bacillus licheniformis, Klebsiella sp., Pantoea dispersa, Klebsiella variicola, Pantoea sp., Agrobacterium larrymoorei and PEEHM-5 endophytic bacterial isolates as Bacillus sp., Bacillus amyloliquefaciens, Lactococcus lactis, Bacillus cereus and Staphylococcus hominis. In planta evaluation of potential isolates at variable chemical fertilizer input indicated their potential in compensating nearly 25% of the fertilizer input as observed on their improvement of shoot and root parameters. Lactococcus lactis inoculation influenced maximum followed by Pantoea and Klebsiella isolates.
The pyrophosphohydrolase RppH is involved in the control of RsmA/CsrA expression in Azotobacter vinelandii and Escherichia coli Microbiol. Res. (IF 2.777) Pub Date : 2018-05-18 Leidy Patricia Bedoya-Pérez, Luis Felipe Muriel-Millán, Soledad Moreno, Elva Quiroz-Rocha, Nancy Rivera-Gómez, Guadalupe Espín
In bacteria, the 5′-end-dependent RNA degradation is triggered by the RNA pyrophosphohydrolase RppH converting tri/diphosphate to monophosphate transcripts. This study shows that in the soil bacterium Azotobacter vinelandii, inactivation of rppH gene negatively affected the production of bioplastic poly-β-hydroxybutyrate (PHB) by reducing the expression at the translational level of PhbR, the specific transcriptional activator of the phbBAC biosynthetic operon. The effect of RppH on the translation of phbR seemed to be exerted through the translational repressor RsmA, as the inactivation of rsmA in the rppH mutant restored the phbR expression. Interestingly, in Escherichia coli inactivation of rppH also affected the expression of CsrA, the RsmA homolog. The level of the csrA transcript was higher and more stable in the E. coli rppH mutant than in the wild type strain. Additionally, and in contrast to the csrA mutants that are known to have a defective swimming phenotype, the E. coli rppH mutant showed a hyper-swimming phenotype that was suppressed by a csrA mutation, and the AvRppH restored to wild type level the swimming phenotype to the E. coli rppH mutant. We propose that in both A. vinelandii and E. coli, RppH activity plays a role in the expression of the translational regulator protein RsmA/CsrA.
Novel Trichoderma strains isolated from tree barks as potential biocontrol agents and biofertilizers for direct seeded rice Microbiol. Res. (IF 2.777) Pub Date : 2018-05-23 Harekrushna Swain, Totan Adak, Arup K. Mukherjee, Prasun K. Mukherjee, Pratap Bhattacharyya, Shantiprava Behera, Torit Baran Bagchi, Rashmishree Patro, Shasmita, Ansuman Khandual, M.K. Bag, T.K. Dangar, S. Lenka, M. Jena
This study is the first time report of utilization of Trichoderma spp. isolated from different tree bark from Odisha state of India for rice crop health management and higher productivity. Six isolates of Trichoderma spp. were identified based on the morphological characteristics and species determination was performed by molecular assays. One of the isolated strains determined as Trichoderma erinaceum outperformed others. Trichoderma erinaceum controlled three soil borne plant pathogens i.e. Rhizoctonia solani, Sclerotium rolfsii and Sclerotium oryzae effectively under controlled condition and R. solani and Helminthosporium oryzae under filed condition. Seed treatments with the formulated isolates improved the germination rate of rice and enhanced vigour. These parameters along with higher chlorophyll content could be related to higher yield observed in two rice varieties; Karuna and Sahabhagidhan. Among the six isolates tested, Trichoderma erinaceum treatment recorded highest yield. Significantly higher expression of some stress related enzymes was observed in Trichoderma treated plants which helped in better crop growth both under biotic and abiotic stresses. These isolates helped both the varieties to accumulate more nutrients. This study proves that Trichoderma erinaceum obtained from tree bark may be incorporated in integrated rice crop management both as biocontrol agent and biofertilizer.
Brazilian red propolis: Chemical composition and antibacterial activity determined using bioguided fractionation Microbiol. Res. (IF 2.777) Pub Date : 2018-05-04 Luciane Corbellini Rufatto, Paola Luchtenberg, Charlene Garcia, Christine Thomassigny, Sylvie Bouttier, João Antonio Pêgas Henriques, Mariana Roesch-Ely, Françoise Dumas, Sidnei Moura
The indiscriminate use of antibiotics is causing an increase in bacterial resistance, complicating therapeutic planning. In this context, natural products have emerged as major providers of bioactive compounds. This work performs a bioguided study of Brazilian red propolis to identify compounds with antibacterial potential and to evaluate their cytotoxicity against non-tumour cells. Using bioguided fractionation performed with the hydroalcoholic extract of red propolis from Alagoas, it was possible to obtain subfractions with remarkable bacteriostatic activity compared with the precursor fractions. The SC2 subfraction was highlighted and showed the best results with minimal inhibitory concentrations (MICs) of 56.75, 28.37, 454.00, and 227.00 μg mL−1 against Staphylococcus aureus, Bacillus subtilis, Escherichia coli, and Pseudomonas aeruginosa, respectively. However, this study also revealed a cytotoxic effect against the non-tumour Vero cell line. Furthermore, through chemical analyses using high resolution mass spectrometry, high performance liquid chromatography with UV detection, and gas chromatography coupled to mass spectrometry, we verified the presence of important marker compounds in the fractions and extracts, including formononetin (m/z 267.0663), biochanin A (m/z 283.0601), and liquiritigenin (m/z 255.0655). The results obtained in this study suggest an important antibacterial potential of red propolis subfractions. In this context, the bioguided fractionation has been a useful process, due to its ability to isolate and concentrate active compounds in a logical and rational way.
LotS/LotR/Clp, a novel signal pathway responding to temperature, modulating protease expression via c-di-GMP mediated manner in Stenotrophomonas maltophilia FF11 Microbiol. Res. (IF 2.777) Pub Date : 2018-05-19 Qingling Wang, Fangling Ji, Jianli Guo, Yuepeng Wang, Yanyan Li, Jingyun Wang, Lijia An, Yongming Bao
Stenotrophomonas maltophilia as one of increasing food spoilage bacteria and fish pathogens has become a threat to aquiculture industry. A major factor contributing to the success of bacterium is its outstanding ability to secrete protease at low temperatures. Here, a cAMP receptor like protein (Clp) shows a positive regulation on this protease, named S. maltophilia temperature-response protease (SmtP). Interestingly, a two-component system, comprising of LotS sensor and LotR regulator, for low-temperature response is also confirmed to modulate SmtP expression with similar effect to Clp. Evidence is presented that LotS/LotR modulates smtP (coding SmtP) expression via Clp: clp promoter activity was reduced significantly at low temperatures and protease activity was partially restored by Clp overexpressed in lotS or lotR deletion strain. Furthermore, as a Clp negative effector, the binding ability of c-di-GMP with Clp is not impacted by temperature. c-di-GMP level was increased in S. maltophilia growing at high temperature, but not exhibited significantly in lotR deleted strain, these indicate that LotR is required for temperature modulating c-di-GMP level, although the synthesis or degradation activity of c-di-GMP by LotR was not detected. These findings suggest that LotS/LotR/Clp play an important role in responding to temperature stimuli via c-di-GMP mediated manner.
Community structure and plant growth-promoting potential of cultivable bacteria isolated from Cameroon soil Microbiol. Res. (IF 2.777) Pub Date : 2018-05-12 Gylaine Vanissa Tchuisseu Tchakounté, Beatrice Berger, Sascha Patz, Henri Fankem, Silke Ruppel
Exploiting native plant growth-promoting rhizobacteria (PGPR) in Cameroonian agro-ecosystems provides a means to improve plant–microbe interactions that may enhance ecosystem sustainability and agricultural productivity in an environmentally eco-friendly way. Consequently, we aimed to investigate the community structure and functional PGPR diversity of maize grown in Cameroon. Native bacteria isolated from Cameroon maize rhizosphere soil were identified by partial 16S rRNA gene sequencing and screened for traits particularly relevant for Cameroon low-fertility soil conditions, such as their abilities to tolerate high concentrations of salt, and their plant growth- promoting potential. Genetic and functional diversity was characterized according to their phylogenetic affiliation. A total of 143 bacteria were identified and assigned to 3 phyla (Actinobacteria, Firmicutes and Proteobacteria), 13 families and 20 genera. Bacillus (31.5%), Arthrobacter (17.5%), and Sinomonas (13.3%) were the most abundant genera identified among all the isolates. Based on their in vitro characterization, 88.1% were salt tolerant at 2% NaCl, but only 16.8% could tolerate 8% NaCl, 50.4% solubilized phosphate, 10.5% possessed the nifH gene, and 19.6% produced siderophores. Six isolates affiliated to the most abundant genera identified in this work, Bacillus and Arthrobacter, carrying multiple or only single tested traits were selected to evaluate their growth- promoting potential in an in vitro maize germination assay. Three strains possessing multiple traits induced significantly increased hypocotyl and root length of maize seeds compared to non-inoculated control seeds. Our results indicate the potential of selected indigenous Cameroon rhizobacteria to enhance maize growth.
Implication of orphan histidine kinase (OhkAsp) in biosynthesis of doxorubicin and daunorubicin in Streptomyces peucetius ATCC 27952 Microbiol. Res. (IF 2.777) Pub Date : 2018-05-22 Anaya Raj Pokhrel, Hue Thi Nguyen, Dipesh Dhakal, Amit Kumar Chaudhary, Jae Kyung Sohng
The orphan histidine kinase (HK) from Streptomyces peucetius ATCC 27952 (ohkAsp) was found to be implicated in the regulation of doxorubicin (DOX)/daunorubicin (DNR) biosynthesis, self-defense and developmental attributes. OhkAsp is a homolog of OhkA from Streptomyces coelicolor and Streptomyces avermitilis (with 73 and 75% identity). As in its homologs, S. peucetius mutant with deletion of ohkAsp was found to enhance metabolite biosynthesis and impaired the morphological differentiation. But, unlike its homologs from Streptomyces coelicolor and Streptomyces avermitilis, differential enhancement in level of secondary metabolite production was found in overexpression mutants apart from deletion mutant. The deflection in characteristics of OhkA in its homologue from S. peucetius ATCC 27952, and its imminent implications was monitered by making various mutants with differential expression level of ohkAsp. The variations were observed in the morphology of mutants, transcriptional level of effectors and regulators of DOX/DNR biosynthesis pathway, DOX/DNR precursor pool and biomass accumulation. Based on comparisons of domain arrangements among its homologs, Low Complexity Region (LCR) present on the OhkAsp was the only domain that stood out. Further, the LCR on OhkAsp was found to be overlapping with a putative receiver domain responsible for interaction with response regulator. The imminent implications of differential expression level of ohkAsp on: regulation and biosynthesis of DOX/DNR, morphological differentiation, DOX/DNR precursor pool and biomass accumulation were explored in this study.
Role of hypothetical protein YicS in the pathogenicity of Avian Pathogenic Escherichia coli in vivo and in vitro Microbiol. Res. (IF 2.777) Pub Date : 2018-05-13 Renu Verma, Thaís Cabrera Galvão Rojas, Renato Pariz Maluta, Janaína Luisa Leite, Gerson Nakazato, Wanderley Dias de Silveira
Avian Pathogenic Escherichia coli (APEC) strains belong to the extra-intestinal pathogenic group of E. coli (ExPEC) that causes colibacillosis in poultry. A variety of putative virulence factors of APEC are recognized as potent causes of pathogenicity, the mechanisms underlying their pathogenicity are still not fully understood. The role of yicS in the virulence of pathogenic E. coli is still unclear. Thus, yicS may be related to biofilm formation, which in some bacteria plays a role in pathogenicity. Therefore, the fact that this gene appears to be under positive selection pressure suggests that yicS may be associated with the pathogenicity of APEC. To better understand the role of yicS protein in APEC biological characteristics and pathogenicity, we deleted yicS in an APEC Swollen Head Syndrome strain (APEC strain SCI-07) and studied its effects by comparing wild type and isogenic mutants through comprehensive in vitro and in vivo assays. We demonstrated that yicS plays a role in pathogenicity of APEC. We suggest that the yicS gene, which encodes an exporter protein, has a significant role in biofilm formation, motility, invasion of CEC-32 and Hep-2 cells and APEC pathogenicity in a day-old chick model.
Symbiotic characteristics of Bradyrhizobium diazoefficiens USDA 110 mutants associated with shrubby sophora (Sophora flavescens) and soybean (Glycine max) Microbiol. Res. (IF 2.777) Pub Date : 2018-05-21 Yuan Hui Liu, En Tao Wang, Yin Shan Jiao, Chang Fu Tian, Lei Wang, Zi Jian Wang, Jia Jing Guan, Raghvendra Pratap Singh, Wen Xin Chen, Wen Feng Chen
Site-specific insertion plasmid pVO155 was used to knockout the genes involved in the alternation of host range of strain Bradyrhizobium diazoefficiens USDA 110 from its original determinate-nodule-forming host soybean (Glycine max), to promiscuous and indeterminate-nodule-forming shrubby legume sophora (Sophora flavescens). Symbiotic phenotypes of these mutants inoculated to these two legumes, were compared to those infected by wild-type strain USDA 110. Six genes of the total fourteen Tn5 transposon mutated genes were broken using the pVO155 plasmid. Both Tn5 and pVO155-inserted mutants could nodulate S. flavescens with different morphologies of low-efficient indeterminate nodules. One to several rod or irregular bacteroids, containing different contents of poly-β-hydroxybutyrate or polyphosphate were found within the symbiosomes in nodulated cells of S. flavescens infected by the pVO155-inserted mutants. Moreover, none of bacteroids were observed in the pseudonodules of S. flavescens, infected by wild-type strain USDA 110. These mutants had the nodulation ability with soybean but the symbiotic efficiency reduced to diverse extents. These findings enlighten the complicated interactions between rhizobia and legumes, i. e., mutation of genes involved in metabolic pathways, transporters, chemotaxis and mobility could alter the rhizobial entry and development of the bacteroid inside the nodules of a new host legume.
Clioquinol induces G2/M cell cycle arrest through the up-regulation of TDH3 in Saccharomyces cerevisiae Microbiol. Res. (IF 2.777) Pub Date : 2018-05-04 Chongjia Yan, Song Wang, Jian Wang, Hui Li, Zhiwei Huang, Jing Sun, Min Peng, Wenbin Liu, Ping Shi
Clioquinol (CQ) has been used as a classical antimicrobial agent for many years. However, its mode of action is still unclear. In our study, the growth of Candida albicans and Saccharomyces cerevisiae was inhibited by CQ. It did not kill yeast cells, but shortened G1 phase and arrested cell cycle at G2/M phase. By using two-dimensional electrophoresis based proteomic approach, six proteins were found to be significantly affected by CQ. Among them, four (PDC1, ADH1, TDH3, IPP1) were up-regulated and the other two (TDH1 and PGK1) were down-regulated. According to the Saccharomyces Genome Database (SGD), these proteins were involved in various biological processes including glycolytic fermentation, gluconeogenesis, glycolytic process, amino acid catabolism, redox reaction and reactive oxygen species metabolic process. It was noted that there was a link between TDH3 and cell cycle. The overexpression of TDH3 phenocopied CQ treatment and arrested the cell cycle at G2/M phase. RT-PCR analysis showed that the mRNA levels of CLN3 and CDC28, critical genes for passage through G1 phase, were up-regulated after the treatment of CQ as well as the overexpression of TDH3. It demonstrates that CQ inhibits the growth of yeast by up-regulating the expression of TDH3 to influence the cell cycle. It is expected to provide new insights for the antimicrobial mechanism of CQ.
Characterization and regulation of AcrABR, a RND-type multidrug efflux system, in Agrobacterium tumefaciens C58 Microbiol. Res. (IF 2.777) Pub Date : 2018-07-10 Puttamas Nuonming, Sasimaporn Khemthong, Thanittra Dokpikul, Rojana Sukchawalit, Skorn Mongkolsuk
Agrobacterium tumefaciens AcrR is the transcriptional repressor of the acrABR operon. The AcrAB efflux pump confers resistance to various toxic compounds, including antibiotics [ciprofloxacin (CIP), nalidixic acid (NAL), novobiocin (NOV) and tetracycline (TET)], a detergent [sodium dodecyl sulfate (SDS)] and a biocide [triclosan (TRI)]. The sequence to which AcrR specifically binds in the acrA promoter region was determined by EMSA and DNase I footprinting. The AcrR-DNA interaction was abolished by adding NAL, SDS and TRI. Quantitative real time-PCR analysis showed that induction of the acrA transcript occurred when wild-type cells were exposed to NAL, SDS and TRI. Indole is a signaling molecule that increases the antibiotic resistance of bacteria, at least in part, through activation of efflux pumps. Expression of the A. tumefaciens acrA transcript was also inducible by indole in a dose-dependent manner. Indole induced protection against CIP, NAL and SDS but enhanced susceptibility to NOV and TRI. Additionally, the TET resistance of A. tumefaciens was not apparently modulated by indole. A. tumefaciens AcrAB played a dominant role and was required for tolerance to high levels of the toxic compounds. Understanding the regulation of multidrug efflux pumps and bacterial adaptive responses to intracellular and extracellular signaling molecules for antibiotic resistance is essential. This information will be useful for the rational design of effective treatments for bacterial infection to overcome possible multidrug-resistant pathogens.
'Ropy' phenotype, exopolysaccharides and metabolism: study on food isolated potential probiotics LAB Microbiol. Res. (IF 2.777) Pub Date : 2018-07-09 Simona Cirrincione, Yael Breuer, Erika Mangiapane, Roberto Mazzoli, Enrica Pessione
Lactic acid bacteria are fully recognized for their industrial applications among which the production and release of exopolysaccharides. In the present investigation, we screened fifteen Lactobacilli in order to find ropy strains, quantify exopolysaccharides and detect proteins specifically associated with the ropy-exopolysaccharide production. The highest ropy-exopolysaccharide producer (L. helveticus 6E8), was grown in stimulating and basal condition (10% and 2% lactose) and subjected to comparative proteomic analysis. The levels of 4 proteins were found significantly increased in the membrane fraction under stimulating conditions: a specific exopolysaccharide biosynthetic protein, a stress-induced protein, a protein involved in secretion and an ATP-synthase subunit. Conversely, several enzymes involved in anabolism and protein synthesis were decreased. These results suggest a general shift from growth to exopolysaccharide-mediated protection from the hyperosmotic environment. Due to the great interest in exopolysaccharides with novel features, the identification of these proteins could have implications for future improvements of industrial strains.
An essential role for hfq involved in biofilm formation and virulence in serotype 4b Listeria monocytogenes Microbiol. Res. (IF 2.777) Pub Date : 2018-07-07 Hao Yao, Meiqin Kang, Yuting Wang, Youwei Feng, Suwei Kong, Xuexue Cai, Zhiting Ling, Sisi Chen, Xin’an Jiao, Yuelan Yin
Regulator factor Hfq has been widely detected among both Gram-positive and Gram-negative bacteria; however, its role in Gram-positive bacteria is less well established and varies among species. In Listeria monocytogenes (Lm), an organism able to adapt to a range of environments and live both saprobiotic and parasitic lifestyles, the role of Hfq is not fully understood. Serotype 4b Listeria monocytogenes strains associated with the majority of listeriosis outbreak, while the function of hfq in serotype 4b strains still not referenced. Here, we constructed hfq deletion and reversion mutants of serotype 4b Lm NTSN and analysed the biological characteristics both in vitro and in vivo. The deletion of hfq resulted in a growth deficiency in medium containing 4.5% ethanol or 1% Triton X-100, and the growth of the mutant was significantly reduced at 4 °C. Furthermore, the hfq deletion dramatically decreased biofilm formation in BHI medium and gastric fluid medium, and reduced the invasion rate into the Caco-2 cell line. However, complementation restored the wild-type phenotype. Importantly, mouse infection experiments demonstrated that hfq played a more important role in the colonisation and virulence in serotype 4b strain Lm NTSN than in the serotype 1/2a strain Lm EGDe. Taken together, these results demonstrated that hfq is a novel factor associated with biofilm formation, and plays an essential role in the stress response and pathogenisis in serotype 4b strain Lm NTSN. Our data provide the basis for further research into the function of Hfq in serotype 4b Listeria monocytogenes.
Time-resolved transcriptome analysis of Clostridium difficile R20291 response to cysteine Microbiol. Res. (IF 2.777) Pub Date : 2018-07-07 Huawei Gu, Kan Shi, Zhengping Liao, Haonan Qi, Shuyi Chen, Haiying Wang, Shan Li, Yi Ma, Jufang Wang
The incidence of Clostridium difficile infection has been steadily rising over the past decade. The increase in the rate of incidence is associated with the specific NAP1/BI/027 strains which are “hypervirulent” and have led to several large outbreaks since their emergence. However, the relation between these outbreaks and virulence regulation mechanisms remains unclear. It has been reported that the major virulence factor TcdA and TcdB in C. difficile could be repressed by cysteine. Here, we investigated the functional and virulence-associated regulation of C. difficile R20291 response to cysteine by using a time-resolved genome-wide transcriptome analysis. Dramatic changes of gene expression in C. difficile revealed functional processes related to transport, metabolism, and regulators in the presence of cysteine during different phases of growth. Flagellar and ribosomal genes were significantly down-regulated in long-term response to cysteine. Many NAP1/BI/027- specific genes were also modulated by cysteine. In addition, cdsB inactivation in C. difficile R20291 could remove the repression of toxin synthesis but could not remove the repression of butyrate production in the presence of cysteine. This suggests that toxin synthesis and butyrate production might have different regulatory controls in response to cysteine. Altogether, our research provides important insights into the regulatory mechanisms of cysteine in C. difficile.
Effectiveness of multi-trait Burkholderia contaminans KNU17BI1 in growth promotion and management of banded leaf and sheath blight in maize seedling Microbiol. Res. (IF 2.777) Pub Date : 2018-05-06 Setu Bazie Tagele, Sang Woo Kim, Hyun Gu Lee, Hyun Seung Kim, Youn Su Lee
Plant growth promoting (PGP) bacteria enhance plant growth and are a green alternative to chemical fertilizers. In our study, an effective plant growth promoting rhizobacteria (PGPR) strain, KNU17BI1, was isolated from rhizospheric soil of maize, South Korea. The strain was tested in vitro for specific PGP and antifungal traits, such as phosphate solubilization, zinc solubilization, indole acetic acid (IAA) production, ammonia production, nitrogen fixation, 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity, siderophore hydrogen cyanide production (HCN) and hydrolytic enzyme activity. Furthermore, in viro antifungal activity was done in a laboratory and in vivo effect of KNU17BI1 on banded leaf and sheath blight intensity as well as plant growth promotion on maize seedling were conducted under greenhouse conditions. The strain was found to be highly effective toward all the parameters except HCN production. The strain KNU17BI1 was identified on the basis of 16S RNA and multilocus sequence analysis (MLSA) and confirmed as Burkholderia contaminans. This study for the first time demonstrated potent in vitro antifungal activity of B. contaminans against Rhizoctonia solani AG-1(IA), Pythium graminicola, Fusarium moniliforme, Alternaria alternata, Alternaria solani, Fusarium graminearum, Stemphylium botryosum Wallr, Colletotrichum dematium, Stemphylium lycopersici and Fusarium oxysporum f.sp. melonis. Furthermore, in this study, for the first time, the potential of B. contaminans stain KNU17BI1 in controlling banded leaf and sheath blight of maize caused by R. solani AG-1(IA) was reported. Therefore, further studies are warranted on the structural identification of actual compounds behind such activities that would be exploited further for biocontrol as well as plant growth promotion.
Fungal communities associated with Evernia prunastri, Ramalina fastigiata and Pleurosticta acetabulum: Three epiphytic lichens potentially active against Candida biofilms Microbiol. Res. (IF 2.777) Pub Date : 2018-03-21 Aurélie Lagarde, Patricia Jargeat, Mélanie Roy, Marion Girardot, Christine Imbert, Marion Millot, Lengo Mambu
Fungal communities associated to three epiphytic lichens active against Candida, were investigated using culture-based methods We hypothetized that associated fungi would contribute to lichens activities. The ability of specific fungi to grow inside or outside lichens was investigated. To detect biogenesis pathways involved in the production of secondary metabolites, genes coding for nonribosomal peptide synthetase (NRPS) and polyketide synthase I (PKS I) were screened by PCR from fungal DNA extracts. Both endo and epilichenic communities were isolated from two fructicose (Evernia prunastri and Ramalina fastigiata) and one foliose (Pleurosticta acetabulum) lichens. A total of 86 endolichenic and 114 epilichenic isolates were obtained, corresponding to 18 and 24 phylogenetic groups respectively suggesting a wide diversity of fungi. The communities and the species richness were distinct between the three lichens which hosted potentially new fungal species. Additionally, the endo- and epilichenic communities differed in their composition: Sordariomycetes were particularly abundant among endolichenic fungi and Dothideomycetes among epilichenic fungi. Only a few fungi colonized both habitats, such as S. fimicola, Cladosporium sp1 and Botrytis cinerea. Interestingly, Nemania serpens (with several genotypes) was the most abundant endolichenic fungus (53% of isolates) and was shared by the three lichens. Finally, 12 out of 36 phylogenetic groups revealed the presence of genes coding for nonribosomal peptide synthetase (NRPs) and polyketide synthase I (PKS I). This study shows that common lichens are reservoirs of diverse fungal communities, which could potentially contribute to global activity of the lichen and, therefore, deserve to be isolated for further chemical studies.
Subterranean infestation by Holotrichia parallela larvae is associated with changes in the peanut (Arachis hypogaea L.) rhizosphere microbiome Microbiol. Res. (IF 2.777) Pub Date : 2018-03-23 Li-Li Geng, Gao-Xiang Shao, Ben Raymond, Mei-Ling Wang, Xiao-Xiao Sun, Chang-Long Shu, Jie Zhang
Rhizosphere microorganisms contribute to the health and development of crops and these beneficial microbes are recruited to the root-zone when plants experience biotic/abiotic stress. The subterranean pests Holotrichia parallela cause severe crop loss in peanut (Arachis hypogaea L.) fields. Hypothesizing that infestation by H. parallela larva may influence the composition of rhizosphere microbial communities, deep sequencing of V3 and V4 hypervariable regions of 16S rRNA gene was used to characterize the rhizosphere bacteria of infested and uninfested peanuts. A total of 2,673,656 reads were generated and an average of 2558 OTUs were obtained for each sample. Comparisons of rhizosphere bacterial community structure of peanuts with those infested by H. parallela larva revealed that the relative abundance of Proteobacteria and Bacteroidetes increased, while that of Actinobacteria decreased in the rhizosphere with infestation. A significant shift in bacterial communities was observed within 24 h after infestation by principal coordinate analysis. For the 332 genera identified in 24 h treatment, infestation of white grubs led to the significant changes of abundance of 67 genera. An increase in the Pseudomonas genus of infested-samples for 24 h was verified by real-time qPCR. Our results indicate H. parallela larvae infestation can quickly leads to the change of peanut rhizosphere microbiome and enrichment of specific bacterial species. But the effects were not persistent. This study provides the insight into the function of rhizosphere microbiome in the interaction between subterranean pests and crops.
The role of efflux pumps in Bacteroides fragilis resistance to antibiotics Microbiol. Res. (IF 2.777) Pub Date : 2018-03-01 Reza Ghotaslou, Mina Yekani, Mohammad Yousef Memar
The resistance of Bacteroides fragilis to the most antimicrobial agents has been reported in the world. Identification of the microbial resistance mechanisms can play an important role in controlling these resistances. Currently, B. fragilis is resistant to most antibiotics. The multi-drug efflux pumps have been shown to underlie the antimicrobial resistance in B. fragilis strains. Two types of these efflux pumps including RND and MATE can be regarded as main structures responsible for antibiotic resistance. Therefore, the strategy for suppressing of this efflux system may be useful in the treatment and control of the multidrug-resistant B. fragilis. The purpose of this study is to review the B. fragilis efflux pumps and their functions in the resistance to antibiotics.
The transcription factor-encoding gene crtf is involved in Clonostachys chloroleuca mycoparasitism on Sclerotinia sclerotiorum Microbiol. Res. (IF 2.777) Pub Date : 2018-03-08 Zhan-Bin Sun, Qi Wang, Jun Zhang, Wei-Zhi Jiang, Qi Wang, Shi-Dong Li, Gui-Zhen Ma, Man-Hong Sun
Clonostachys chloroleuca 67-1 (formerly C. rosea 67-1) is a potential biocontrol fungus active against various fungal plant pathogens. From transcriptome sequencing of 67-1 parasitizing sclerotia of Sclerotinia sclerotiorum, we identified the transcription factor-encoding gene crtf that is significantly up-regulated during mycoparasitism. Transcription factors are widely distributed in fungi and involved in multiple biological processes. However, their role and regulatory mechanisms in mycoparasitism remain poorly understood. In this study, the function of crtf during 67-1 mycoparasitism was verified through gene knockout and complementation. The results showed that deletion of crtf did not influence fungal morphological characteristics, but the ability of the Δcrtf mutant to parasitize sclerotia and suppress soybean Sclerotinia white mold in the greenhouse was markedly diminished compared with the wild type strain. The biocontrol activity of Δcrtf recovered wild type levels when complemented with a plasmid expressing the crtf gene. These findings suggest that crtf plays a crucial role in C. chloroleuca mycoparasitism and provide insight into the molecular mechanisms underlying C. chloroleuca mycoparasitism on plant pathogenic fungi.
Characterization of Cd-resistant Klebsiella michiganensis MCC3089 and its potential for rice seedling growth promotion under Cd stress Microbiol. Res. (IF 2.777) Pub Date : 2018-03-08 Soumik Mitra, Krishnendu Pramanik, Pallab Kumar Ghosh, Tithi Soren, Anumita Sarkar, Ramendra Sundar Dey, Sanjeev Pandey, Tushar Kanti Maiti
Application of heavy metal resistant plant growth promoting rhizobacteria has an important role as they help to evade metal-induced toxicity in plants on one hand and enhance plant growth on the other. The present study is therefore focused on the characterization of a cadmium resistant bacterial strain isolated from heavy metal contaminated rhizospheric soil designated as S8. This S8 strain was selected in terms of cadmium resistance and plant growth promoting traits. Moreover, it also showed resistance to lead and arsenic to a considerable extent. The selected strain S8 was identified as Klebsiella michiganensis by modern approaches of bacterial taxonomy. The plant growth promoting traits exhibited by the strain include 1-aminocyclopropane-1-carboxylic acid deaminase activity (58.33 ng α-keto butyrate/mg protein/h), Indole-3-acetic acid production (671 μg/ml), phosphate solubilization (71.98 ppm), nitrogen fixation (3.72 μg of nitrogen fixed/h/mg protein) etc. Besides, the strain also exhibited high cadmium removal efficiency (73–97%) from the medium and intracellular accumulation as well. Its efficiency to alleviate cadmium-induced toxicity was determined against a rice cultivar in terms of morphological and biochemical changes. Enhanced growth and reduced oxidative stress were detected in presence of the bacterium. On the basis of these results, it can be concluded that K. michiganensis strain S8 is cadmium accumulating plant growth promoting rhizobacterium that can be applied in cadmium contaminated agricultural soil to achieve better productivity of rice.
Identification and characterization of a new Bacillus atrophaeus strain B5 as biocontrol agent of postharvest anthracnose disease in soursop (Annona muricata) and avocado (Persea americana) Microbiol. Res. (IF 2.777) Pub Date : 2018-02-02 Lizeth Guardado-Valdivia, Erik Tovar-Pérez, Alejandra Chacón-López, Ulises López-García, Porfirio Gutiérrez-Martínez, Alexandra Stoll, Selene Aguilera
Anthracnose is a fungal disease caused by Colletotrichum species that is detrimental to numerous fruit, including soursop and avocado. The use of fungicides to maintain the high quality of fruit creates a potential health risk. One alternative to this problem is the biological control, which has been applied successfully during postharvest. The Bacillus species are one of the most studied biological agents against postharvest pathogens because accomplish their biocontrol performance by producing a variety of metabolites. In this study, we evaluated the activity of metabolites contained in the cell free supernatant, obtained from Bacillus strain B5 culture, against micelial growth and spore germination of two virulent strains of C. gloeosporioides isolated from soursop and avocado. On the basis of 16S rDNA gene sequence analysis, this strain was identified as Bacillus atrophaeus. A preventive treatment using cell free supernatant, reduced severity and incidence of anthracnose disease on harvested soursop and avocado fruit. B. atrophaeus strain B5 harbors genes involved in the production of antibiotics such as surfactin, bacillomycin and iturin, which could be contributing to the efficiency of the preventive treatment during postharvest. The antagonistic role of metabolites contained in the cell free supernatant against anthracnose disease, provide a new approach by which to attack this problem and can help reduce the use of chemical pesticides, environmental pollution, leading to the safer fruit preservation.
Roles of phospholipid methyltransferases in pycnidia development, stress tolerance and secondary metabolism in the taxol-producing fungus Pestalotiopsis microspore Microbiol. Res. (IF 2.777) Pub Date : 2018-03-08 Oren Akhberdi, Qian Zhang, Haichuan Wang, Yingying Li, Longfei Chen, Dan Wang, Xi Yu, Dongsheng Wei, Xudong Zhu
Phosphatidylcholine (PC) is an important membrane component of the eukaryotic cell. In yeast fungi, two phospholipid methyltransferases catalyze consecutive steps of methylation in the formation of phosphatidylcholine from phosphatidylethanolamine. However, roles of phospholipid methyltransferases in filamentous fungi remains less investigated. We report here the characterization of two genes, choA and choC, that putatively encoded phospholipid methyltransferases in the taxol-producing fungus Pestalotiopsis microspora. Deletion of choC resulted in defects in PC production, vegetative growth and development of asexual structure. The mutant strains exhibited multiple morphological abnormalities, e.g. swollen hyphal tips and enhanced hyphal branching, and even mycelial autolysis. Some novel roles for the genes were also revealed, for instance, the deletion of either choC or choA impaired the development of pycnidia and conidia, the cell wall integrity. The mutant strains displayed a hypersensitivity to stress conditions, e.g. osmotic stress, cold and metal ions. The osmotic hypersensitivity indicates a crosstalk of PC pathways to other signaling pathways, such as the HOG pathway. Still more, choA, but not choC, was required for the production of secondary metabolites, e.g. pestalotiollide B, suggesting distinct roles of the two genes. This work would contribute to better understanding the function of phospholipid methyltransferases in fungi.
Multicellular behavior of environmental Escherichia coli isolates grown under nutrient-poor and low-temperature conditions Microbiol. Res. (IF 2.777) Pub Date : 2018-03-15 Laura Di Sante, Armanda Pugnaloni, Francesca Biavasco, Eleonora Giovanetti, Carla Vignaroli
The multicellular behavior designated “red dry and rough” (rdar) morphotype—characterized by production of extracellular matrix mainly comprising curli fimbriae and cellulose—is a potential survival strategy of Escherichia coli outside the host. This study documents the ability of Escherichia cryptic clades, which have recently been recognized as new lineages genetically divergent from E. coli, to grow in unfavorable conditions through expression of distinct phenotypes. Growth under low-temperature and nutrient-poor conditions induced the rdar morphotype in all cryptic clade strains tested, especially after preincubation in broth supplemented with uracil. Such phenotypic response to harsh growth conditions was clearly detected by transmission and scanning electron microscopy, which showed that bacteria were encased in a fibrous matrix. Conversely, cells incubated in rich medium at 37 °C showed no matrix. Uracil enhanced the biosynthesis of matrix components, fostering biofilm production and strain adhesion to abiotic surfaces, as demonstrated by the increase of strong biofilm producers in biofilm assays. Harsh growth conditions also induced catalase activity, resulting in clade strain resistance to hydrogen peroxide oxidative stress. The present findings further support the ‘environmental hypothesis’ whereby cryptic clades would be able to persist in natural habitats outside the host through the expression of distinct survival phenotypes.
Quorum sensing: A less known mode of communication among fungi Microbiol. Res. (IF 2.777) Pub Date : 2018-03-21 Sajad Ahmad Padder, Rajendra Prasad, Abdul Haseeb Shah
Quorum sensing (QS), a density-dependent signaling mechanism of microbial cells, involves an exchange and sense of low molecular weight signaling compounds called autoinducers. With the increase in population density, the autoinducers accumulate in the extracellular environment and once their concentration reaches a threshold, many genes are either expressed or repressed. This cell density-dependent signaling mechanism enables single cells to behave as multicellular organisms and regulates different microbial behaviors like morphogenesis, pathogenesis, competence, biofilm formation, bioluminescence, etc guided by environmental cues. Initially, QS was regarded to be a specialized system of certain bacteria. The discovery of filamentation control in pathogenic polymorphic fungus Candida albicans by farnesol revealed the phenomenon of QS in fungi as well. Pathogenic microorganisms primarily regulate the expression of virulence genes using QS systems. The indirect role of QS in the emergence of multiple drug resistance (MDR) in microbial pathogens necessitates the finding of alternative antimicrobial therapies that target QS and inhibit the same. A related phenomenon of quorum sensing inhibition (QSI) performed by small inhibitor molecules called quorum sensing inhibitors (QSIs) has an ability for efficient reduction of gene expression regulated by quorum sensing. In the present review, recent advancements in the study of different fungal quorum sensing molecules (QSMs) and quorum sensing inhibitors (QSIs) of fungal origin along with their mechanism of action and/or role/s are discussed.
Inhibition of coxsackievirus B4 by Lactobacillus plantarum Microbiol. Res. (IF 2.777) Pub Date : 2018-03-19 Mattia Pia Arena, Firas Elmastour, Famara Sane, Djamel Drider, Daniela Fiocco, Giuseppe Spano, Didier Hober
The enterovirus Coxsackievirus B4 (CV-B4) can infect different human tissues and provoke abnormal function or destruction of various organs and cells. Moreover, its infections have been linked to the onset of type 1 diabetes. Coxsackievirus B4 is classified as a “challenging virus”, due to the intense yet vain efforts to find effective prevention and therapeutic agents, especially within biological compounds. Lactobacillus plantarum is a lactic acid bacterium that is endowed with probiotic properties, and holds great potential for applications in medical and food industry sectors. Several compounds produced by this microorganism have been associated with various benefits including antimicrobial activity. In this work, we investigated the possible antiviral abilities of two Lb. plantarum strains and their derivatives against CV-B4. The different assays carried out (e.g. pre-incubation, competition and post-infection, using HEp-2 cells as human cell model) suggest that the tested microorganisms and their derivatives have an in vitro inhibiting activity against CV-B4. This is the first report showing the anti-CVB4 activity of Lb. plantarum strains and their derivatives.
Antibiotic-producing Pseudomonas fluorescens mediates rhizome rot disease resistance and promotes plant growth in turmeric plants Microbiol. Res. (IF 2.777) Pub Date : 2018-03-27 S.R. Prabhukarthikeyan, U. Keerthana, T. Raguchander
Rhizome rot of turmeric caused by Pythium aphanidermatum is a major threat to turmeric-cultivating areas of India. This study intends to evaluate the performance of fluorescent pseudomonads against Rhizome rot disease and understand the resistance mechanism in Turmeric plants. Fluorescent pseudomonads were screened against Pythium aphanidermatum using dual culture. Selected strains were evaluated for the performance of growth promoting attributes and the presence of antibiotic genes through PCR analysis. Strain FP7 recorded the maximum percent inhibition of P. aphanidermatum under in vitro conditions. Strains FP7 and TPF54 both increased plant growth in turmeric plants in vitro. Strain FP7 alone contained all the evaluated antibiotic biosynthetic genes. Talc and liquid-based formulations were prepared with effective strain and tested for its biocontrol activities under both glasshouse and field conditions. Enzymatic activities of the induced defense enzymes such as PO, PPO, PAL, CAT and SOD were estimated and subjected to spectrophotometric analysis. A combination of rhizome dip and soil drench of FP7 liquid formulation treatment remarkably recorded the minimum disease incidence, higher defense enzymes, maximum plant growth and yield under glasshouse and field conditions. Application of strain FP7 increased the defense molecules, plant growth and yield in turmeric plants thereby reducing the incidence of rhizome rot disease. Moreover, this study has a potential to be adopted for sustainable and eco-friendly turmeric production.
Biocontrol activity of surfactin A purified from Bacillus NH-100 and NH-217 against rice bakanae disease Microbiol. Res. (IF 2.777) Pub Date : 2018-02-15 Ambrin Sarwar, Muhammad Nadeem Hassan, Muhammad Imran, Mazhar Iqbal, Saima Majeed, Günter Brader, Angela Sessitsch, Fauzia Yusuf Hafeez
The potential of the Bacillus genus to antagonize phytopathogens is associated with the production of cyclic lipopeptides. Depending upon the type of lipopeptide, they may serve as biocontrol agents that are eco-friendly alternatives to chemical fertilizers. This study evaluates the biocontrol activity of surfactin-producing Bacillus (SPB) strains NH-100 and NH-217 and purified surfactin A from these strains against rice bakanae disease. Biologically active surfactin fractions were purified by HPLC, and surfactin A variants with chain lengths from C12 to C16 were confirmed by LCMS-ESI. In hemolytic assays, a positive correlation between surfactin A production and halo zone formation was observed. The purified surfactin A had strong antifungal activity against Fusarium oxysporum, F. moniliforme, F. solani, Trichoderma atroviride and T. reesei. Maximum fungal growth suppression (84%) was recorded at 2000 ppm against F. moniliforme. Surfactin A retained antifungal activity at different pH levels (5–9) and temperatures (20, 50 and 121 °C). Hydroponic and pot experiments were conducted to determine the biocontrol activity of SPB strains and the purified surfactin A from these strains on Super Basmati rice. Surfactin production in the rice rhizosphere was detected by LCMS-ESI at early growth stages in hydroponics experiments inoculated with SPB strains. However, the maximum yield was observed with a consortium of SPB strains (T4) and purified surfactin A (T5) treatments in the pot experiment. The outcomes of the present study revealed that surfactin A significantly reduced rice bakanae disease by up to 80%. These findings suggest that purified surfactin A could be an effective biocontrol agent against bakanae disease in rice and should be incorporated into strategies for disease management.
Transcriptional control of the phenol hydroxylase gene phe of Corynebacterium glutamicum by the AraC-type regulator PheR Microbiol. Res. (IF 2.777) Pub Date : 2018-02-06 Can Chen, Yaoling Zhang, Lei Xu, Kaixiang Zhu, Yanyan Feng, Junfeng Pan, Meiru Si, Lei Zhang, Xihui Shen
Corynebacterium glutamicum can degrade phenol by a meta-cleavage pathway, which depends on ncgl2588 (phe) of the phe operon encoding phenol hydroxylase. An additional gene, ncgl2587 (pheR), is located upstream of phe. The pheR encodes an AraC/XylR-type regulator protein with 377 amino acid residues and is transcribed in the same direction as phe. Disruption of pheR by homologous recombination resulted in the accumulation of phenol in C. glutamicum. PheR demonstrates a low type of constitutive expression where phenol induces phe expression. PheR shares 75% sequence identity with AraC-type regulator of Corynebacterium lubricantis and 37 conserved residues, characteristic of AraC family, were located. A constructed pK18mobsacB-Pphe:lacZ transcriptional fusion plasmid was transformed into the wild-type, ΔpheR, and ΔpheR+ strains, and the results indicated that PheR activates the expression of phe encoding phenol hydroxylase. Electrophoretic mobility shift assay (EMSA) demonstrated a direct interaction of PheR with the phe promoter region and binding site of PheR on the Pphe was located 109-bp upstream of phe, as indicated by foot printing analysis. Our research provides deep insight into PheR expression and its regulatory function on Phe in C. glutamicum.
Plant growth promoting bacteria as an alternative strategy for salt tolerance in plants: A review Microbiol. Res. (IF 2.777) Pub Date : 2018-02-13 Muhammad Numan, Samina Bashir, Yasmin Khan, Roqayya Mumtaz, Zabta Khan Shinwari, Abdul Latif Khan, Ajmal Khan, Ahmed AL-Harrasi
Physiologically distinct subpopulations formed in Escherichia coli cultures in response to heat shock Microbiol. Res. (IF 2.777) Pub Date : 2018-02-13 Bożena Bruhn-Olszewska, Paweł Szczepaniak, Ewelina Matuszewska, Dorota Kuczyńska-Wiśnik, Karolina Stojowska-Swędrzyńska, María Moruno Algara, Ewa Laskowska
Bacteria can form heterogeneous populations containing phenotypic variants of genetically identical cells. The heterogeneity of populations can be considered a bet-hedging strategy allowing adaptation to unknown environmental changes – at least some individual subpopulations or cells might be able to withstand future adverse conditions. Using Percoll gradient centrifugation, we demonstrated that in an Escherichia coli culture exposed to heat shock at 50 °C, two physiologically distinct subpopulations were formed. A high-density subpopulation (HD50) demonstrated continued growth immediately after its transfer to LB medium, whereas the growth of a low-density subpopulation (LD50) was considerably postponed. The LD50 subpopulation contained mainly viable but non-culturable bacteria and exhibited higher tolerance to sublethal concentrations of antibiotics or H2O2 than HD50 cells. The levels of aggregated proteins and main molecular chaperones were comparable in both subpopulations; however, a decreased number of ribosomes and a significant increase in protein oxidation were observed in the LD50 subpopulation as compared with the HD50 subpopulation. Interestingly, under anaerobic heat stress, the formation of the HD50 subpopulation was decreased and culturability of the LD50 subpopulation was significantly increased. In both subpopulations the level of protein aggregates formed under anaerobic and aerobic heat stress was comparable. We concluded that the formation of protein aggregates was independent of oxidative damage induced by heat stress, and that oxidative stress and not protein aggregation limited growth and caused loss of LD50 culturability. Our results indicate that heat stress induces the formation of distinct subpopulations differing in their ability to grow under standard and stress conditions.
Heat stress-induced reactive oxygen species participate in the regulation of HSP expression, hyphal branching and ganoderic acid biosynthesis in Ganoderma lucidum Microbiol. Res. (IF 2.777) Pub Date : 2018-02-19 Rui Liu, Xue Zhang, Ang Ren, Deng-Ke Shi, Liang Shi, Jing Zhu, Han-Shou Yu, Ming-Wen Zhao
Heat stress (HS) is an important environmental factor that affects the growth and metabolism of edible fungi, but the molecular mechanism of the heat stress response (HSR) remains unclear. We previously reported that HS treatment increased the length between two hyphal branches and induced the accumulation of ganoderic acid biosynthesis and the gene expression of heat shock proteins (HSPs) in Ganoderma lucidum. In this study, we found that HS induced a significant increase in the cytosolic ROS concentration, and exogenously added ROS scavengers NAC, VC and NADPH oxidase (Nox) inhibitor DPI reduce the cytosolic ROS accumulation in G. lucidum. In addition, the phenomena of the increased gene expression and increased length between the two hyphal branches and the accumulation of GA biosynthesis induced by HS were mitigated. Furthermore, we investigated the effects of HS on Nox-silenced strains (NoxABi-10, NoxABi-11 and NoxRi-4, NoxRi-7) and found that the level of ROS concentration was lower than that in wild-type (WT) strains treated with HS. Additionally, Nox silenced strains reduced the HS-induced increase in HSP expression, the length between two hyphal branches and GA biosynthesis compared with the WT strain. These data indicate that HS-induced ROS participate in the regulation of HSP expression, hyphal branching and ganoderic acid biosynthesis in G. lucidum. In addition, these findings identified potential pathways linking ROS networks to HSR, physiological and metabolic processes in fungi and provide a valuable reference for studying the role of ROS in HSR, mycelium growth and secondary metabolites.
Phospholipases play multiple cellular roles including growth, stress tolerance, sexual development, and virulence in fungi Microbiol. Res. (IF 2.777) Pub Date : 2018-01-03 Ananya Barman, Dibakar Gohain, Utpal Bora, Ranjan Tamuli
Phospholipases are ubiquitous enzymes that hydrolyze phospholipids. Based on the cleavage site of the ester linkage in the substrate phospholipids, phospholipases are classified into four major types, phospholipase A (PLA), phospholipase B (PLB), phospholipase C (PLC), and phospholipase D (PLD), which are further classified into various subtypes. Phospholipases hydrolyze phospholipids into various signaling products including phosphatidic acid (PA), diacylglycerol (DAG), free fatty acids (FFAs), and lyso-phospholipids (LPLs). These signaling products regulate numerous processes such as cytoskeletal dynamics, growth, homeostasis, membrane remodeling, nutrient acquisition, secretion, signal transduction, stress tolerance, sexual development, and virulence in various organisms including fungi. Due to these key cellular roles, phospholipases are also promising targets in diagnostic and therapeutic applications. In this review, we discuss current knowledge about the cellular roles of different classes of phospholipases in fungi.
Some contents have been Reproduced by permission of The Royal Society of Chemistry.
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