Response surface methodology mediated optimization of Lignin peroxidase from Bacillus mycoides isolated from Simlipal Biosphere Reserve, Odisha, India

Background Lignin is a complex polymer of phenyl propanoid units found in the vascular tissues of the plants as one of lignocellulose materials. Many bacteria secrete enzymes to lyse lignin, which can be essential to ease the production of bioethanol. Current research focused on the study of ligninolytic bacteria capable of producing lignin peroxidase (LiP) which can help in lignin biodegradation and bioethanol production. Ligninolytic bacterial strains were isolated and screened from the soil samples of Simlipal Biosphere Reserve (SBR), Odisha (India), for the determination of their LiP activity. Enzymatic assay and optimization for the LiP activity were performed with the most potent bacterial strain. The strain was identified by morphological, biochemical, and molecular methods. Results In this study, a total of 16 bacteria (Simlipal ligninolytic bacteria [SLB] 1–16) were isolated from forest soils of SBR using minimal salt medium containing lignin. Out of the 16 isolates, 9 isolates showed decolourization of methylene blue dye on LB agar plates. The bacterial isolates such as SLB8, SLB9, and SLB10 were able to decolourize lignin with 15.51%, 16.80%, and 33.02%, respectively. Further enzyme assay was performed using H2O2 as substrate and methylene blue as an indicator for these three bacterial strains in lignin containing minimal salt medium where the isolate SLB10 showed the highest LiP activity (31.711 U/mg). The most potent strain, SLB10, was optimized for enhanced LiP enzyme activity using response surface methodology. In the optimized condition of pH 10.5, temperature 30 °C, H2O2 concentration 0.115 mM, and time 42 h, SLB10 showed a maximum LiP activity of 55.947 U/mg with an increase of 1.76 times from un-optimized condition. Further chemical optimization was performed, and maximum LiP activity as well as significant dye-decolourization efficiency of SLB10 has been found in bacterial growth medium supplemented individually with cellulose, yeast extract, and MnSO4. Most notably, yeast extract and MnSO4-supplemented bacterial culture medium were shown to have even higher percentage of dye decolourization compared to normal basal medium. The bacterial strain SLB10 was identified as Bacillus mycoides according to morphological, biochemical, and molecular (16S rRNA sequencing) characterization and phylogenetic tree analysis. Conclusion Result from the present study revealed the potential of Bacillus mycoides bacterium isolated from the forest soil of SBR in producing LiP enzyme that can be evaluated further for application in lignin biodegradation and bioethanol production. Scaling up of LiP production from this potent bacterial strain could be useful in different industrial applications. Graphical Abstract

Isolation and determination of bacterial microbiota of Varroa destructor and isolation of Lysinibacillus sp. from it

Background The importance of bees for environmental health is known. Within the scope of this importance, it is of great importance to protect the health of bees and to prevent colony extinction. In this context, it is very important to develop effective methods in combating microorganisms, parasitoids, mites and organisms that cause disease or harm in bees. Both use different methods in terms of bee health. Result In this study, the possibility of the bacteria isolated from Varroa destructor mite being bioinsecticide was investigated. Accordingly, six bacteria were isolated from the mite. Isolated bacteria were analyzed according to biochemical tests, molecular analysis, optimum growth pH and phylogenetic tree drawn as Pantoea dispersa (GV1), Lysinibacillus macroides (GV3), Bacillus mycoides (GV4), Lysinibacillus fusiformis (GV5), Pseodomonas lutea (GV5), Lysinibacillus varians (GV7). Lysinibacillus sp. The entomopathogenic feature of Lysinibacillus sp. ranked it as the most important species. When the insecticidal properties of bacteria were examined, they were determined as 53, 90, 62, 95, 74 and 83% for GV1, GV3, GV4, GV5, GV6 and GV7, respectively. Conclusion Based on these results and literature review, Lysinibacillus sp. species had a high potential to be used as bioinsecticide against V. destructor mite.

Impact of Algicidal Bacillus mycoides on Diatom Ulnaria acus from Lake Baikal

Algae–bacteria interactions play an important role in water ecosystems. In this work, the BS2-15 algicidal strain was isolated from the bottom sediments of Lake Baikal and identified as Bacillus mycoides on the basis of 16S rDNA sequencing, its described ultrastructure, and biochemical properties. B. mycoides BS2-15 was demonstrated to have a strong algicidal effect against a freshwater diatom culture of Ulnaria acus, inhibiting its growth and increasing frustules fragility. By analyzing the impact of bacterial filtrate onto the cells of U. acus, we demonstrated that perhaps an algicidal compound is produced by bacteria independently in the presence of diatoms in a medium. Using methods of TUNEL and confocal microscopy, we revealed that the bacterial algicidal effect on the diatom cells results in DNA fragmentation, nucleus destruction, and neutral lipid accumulation. This phenomenon highlights the complexity of algae–bacteria interactions and their potential role in regulating water ecosystem microbial populations.

Untargeted Metabolomics Investigation on Selenite Reduction to Elemental Selenium by Bacillus mycoides SeITE01

Bacillus mycoides SeITE01 is an environmental isolate that transforms the oxyanion selenite (SeO32−) into the less bioavailable elemental selenium (Se0) forming biogenic selenium nanoparticles (Bio-SeNPs). In the present study, the reduction of sodium selenite (Na2SeO3) by SeITE01 strain and the effect of SeO32− exposure on the bacterial cells was examined through untargeted metabolomics. A time-course approach was used to monitor both cell pellet and cell free spent medium (referred as intracellular and extracellular, respectively) metabolites in SeITE01 cells treated or not with SeO32−. The results show substantial biochemical changes in SeITE01 cells when exposed to SeO32−. The initial uptake of SeO32− by SeITE01 cells (3h after inoculation) shows both an increase in intracellular levels of 4-hydroxybenzoate and indole-3-acetic acid, and an extracellular accumulation of guanosine, which are metabolites involved in general stress response adapting strategies. Proactive and defensive mechanisms against SeO32− are observed between the end of lag (12h) and beginning of exponential (18h) phases. Glutathione and N-acetyl-L-cysteine are thiol compounds that would be mainly involved in Painter-type reaction for the reduction and detoxification of SeO32− to Se0. In these growth stages, thiol metabolites perform a dual role, both acting against the toxic and harmful presence of the oxyanion and as substrate or reducing sources to scavenge ROS production. Moreover, detection of the amino acids L-threonine and ornithine suggests changes in membrane lipids. Starting from stationary phase (24 and 48h), metabolites related to the formation and release of SeNPs in the extracellular environment begin to be observed. 5-hydroxyindole acetate, D-[+]-glucosamine, 4-methyl-2-oxo pentanoic acid, and ethanolamine phosphate may represent signaling strategies following SeNPs release from the cytoplasmic compartment, with consequent damage to SeITE01 cell membranes. This is also accompanied by intracellular accumulation of trans-4-hydroxyproline and L-proline, which likely represent osmoprotectant activity. The identification of these metabolites suggests the activation of signaling strategies that would protect the bacterial cells from SeO32− toxicity while it is converting into SeNPs.

Pengaruh Pemberian Plant Growth-Promoting Bacteria Indigenous terhadap Pertumbuhan Tanaman Bawang Merah (Allium ascalonicum)

Penelitian Plant Growth-Promoting Bacteria (PGPB) banyak dikembangkan untuk menerapkan sistem pertanian yang berkelanjutan. Hasil dari eksplorasi PGPB indigenous di alam seperti UB Forest menambah informasi pengendalian hayati yang ramah lingkungan. Tujuan dari penelitian ini adalah untuk mengetahui isolat PGPB indigenous yang diisolasi dari serasah kopi UB Forest memiliki pengaruh terhadap pertumbuhan tanaman bawang merah. Penelitian ini dilakukan pada Februari – April 2020 di KabupatenMalang menggunakan Rancangan Acak Kelompok (RAK) enam perlakuan dan tiga ulangan. Penelitian menggunakan seed treatment pada bibit bawang merah sehat varietas Philip tanpa perlakuan inokulasi patogen. Perlakuan yang digunakan antara lain kontrol dan lima isolat PGPB indigenous yaitu Alcaligenes faecalis, Bacillus mycoides, Clostridium sp., Erwinia sp., dan Pseudomonas sp.Hasil penelitian menunjukkan bahwa pemberian PGPB indigenous memberikan pengaruh yang nyata pada pertumbuhan tinggi tanaman bawang merah. Namun, peningkatan parameter pertumbuhan tersebut tidak diikuti oleh peningkatan jumlah daun dan produksi senyawa ketahanan yaitu enzim peroksida yang dihasilkan pada daun bawang merah. Penelitian ini merupakan penelitian awal untuk menentukan isolat PGPB indigenous yang selanjutnya akan diuji kemampuannya dalam meningkatkan pertumbuhan tanaman bawang merah apabila dilakukan induksi penyakit layu fusarium.

Pan-Genome Portrait of Bacillus mycoides Provides Insights into the Species Ecology and Evolution

This research allows deeper understanding of the genetic organization of natural bacterial populations, specifically, Bacillus mycoides , a psychrotrophic member of the Bacillus cereus group that is widely distributed worldwide, especially in areas with continental cold climates. These thorough analyses made it possible to describe, for the first time, the B. mycoides pan-genome, phylogenetic relationship within this species, and the mechanisms behind the species ecology and evolutionary history.

PEMANFAATAN BIODIVERSITAS BAKTERI SERASAH KOPI SEBAGAI SOLUSI PENGENDALI PENYAKIT MOLER PADA BAWANG MERAH

Penyakit busuk pangkal batang/moler (Fusarium oxysporum f.sp. cepae/FOC) merupakan salah satu faktor biotik dalam produksi bawang merah yang menyebabkan kerusakan hingga 50 %. Pengendalian biologi menawarkan strategi potensial pada perlindungan tanaman yang berfokus pada peran musuh alami dalam menekan hama dan penyakit tanaman. Penelitian ini bertujuan untuk mengidentifikasi dan mengetahui jenis bakteri yang berpotensi sebagai agens hayati terhadap penyakit moler pada bawang merah. Penelitian dilaksanakan di kondisi laboratorium dan lapang menggunakan rancangan acak kelompok untuk membandingkan setiap isolat dalam mengendalikan patogen. Sebanyak 5 dari 21 isolat bakteri menunjukkan sifat antagonisme yang tinggi terhadap FOC. Kelima isolat teridentifikasi sebagai Bacillus mycoides (K1), Clostridium sp. (K2), Pseudomonas sp. (K11), Erwinia sp. (K13), dan Pseudomonas sp. (K29). Pseudomonas sp menujukkan aktivitas penghambatan jamur sebesar 61,01 % dan 73,05 % di laboratorium, sedangkan Bacillus mycoides memberikan hasil terbaik pada kondisi lapang. K2 dan K29 menunjukkan hasil terbaik dalam meningkatkan tinggi tanaman dibandingkan dengan kontrol. Sementara itu, K29 juga berpotensi meningkatkan jumlah daun dan berat basah tanaman.