Strain S1 Research Articles

The Complete Genome Sequence of Sclerotinia sclerotiorum (S1), One of the Pathogens Causing Sclerotinosis in Mulberry Fruit

Sclerotinia sclerotiorum is a fungal pathogen that infects an extremely wide range of hosts and can cause diseases in over 600 plant species. In this study, Sclerotinia sclerotiorum (strain S1), which causes sclerotinosis in mulberry fruits, was isolated from the mulberry orchard at the Chongqing Academy of Agricultural Sciences, China, and its complete genome was sequenced using PacBio Sequel II. The assembly consists of 24 contigs with an estimated genome size of 38.91 Mbp and is predicted to encode 9,920 protein-coding genes. This is the first genomic information for Sclerotinia sclerotiorum isolated from diseased mulberry fruit and represents a valuable resource for a comprehensive understanding of Sclerotinia spp. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .

The silver lining of antibiotic resistance: Bacterial-mediated reduction of tetracycline plant stress via antibiotrophy

The reuse of water using effluents containing antibiotics from anthropogenic activities has been mainly linked to the development of antibiotic resistance. However, we report that the development of bacterial tolerance promotes plant growth. In the present study, we aimed to evaluate the efficiency of inoculation of a new antibiotic-degrading bacterium, Erwinia strain S9, in augmenting the tolerance of pea (Pisum sativum L.) plants to tetracycline (TET) (10 and 20 mg/L). Physiological parameters such as tissue elongation and biomass, as well as relative water content, were remarkably lower in plants exposed to TET than in the control. The inhibitory effects of TET were associated with reduced CO2 assimilation, stomatal conductance, transpiration, dark respiration, and light saturation point (LSP). High concentrations of TET-induced oxidative stress are attested by the overproduction of superoxide radicals (O2•−), hydrogen peroxide (H2O2), and hydroxyl radicals (HO•), resulting in increased malondialdehyde content and cell death. The high activity of antioxidant enzymes such as catalase, ascorbate peroxidase, and guaiacol peroxidase validated the proposed mechanism. Under TET stress conditions, supplementation with Erwinia strain S9 was beneficial to pea plants through osmotic adjustment, increased nutrient uptake, gas exchange optimization, and increased antioxidant activities. Its presence not only ensures plant survival and growth during antibiotic stress but also degrades TET via significant antibiotrophy. This strategy is a cost-effective environmental chemical engineering tool that can be used to depollute wastewater or to improve crop resistance in rhizofiltration treatment when treated wastewater is reused for irrigation.

Transcriptional control of carbohydrate catabolism by the CcpA protein in the ruminal bacterium Streptococcus bovis.

As a potent, pleiotropic regulatory protein in Gram-positive bacteria, catabolite control protein A (CcpA) mediates the transcriptional control of carbohydrate metabolism in Streptococcus bovis, a lactate-producing bacterium that plays an essential role in rumen acidosis in dairy cows. Although the rumen uptake of carbohydrates is multi-substrate, the focus of S. bovis research thus far has been on the glucose. With the aid of gene deletion, whole-genome sequencing, and transcriptomics, we have unraveled the role of CcpA in carbohydrate metabolism, on the one hand, and acidosis, on the other, and we show that the S. bovis strain S1 encodes "Carbohydrate-Active Enzymes" and that ccpA deletion slows the organism's growth rate and modulates the organic acid fermentation pathways toward lower lactate, higher formate, and acetate in the maltose and cellobiose. Furthermore, this study revealed the different regulatory functions of the CcpA protein in rumen metabolism and acidosis.IMPORTANCEThis study is important as it illustrates the varying regulatory role of the Streptococcus bovis catabolite control protein A protein in carbohydrate metabolism and the onset of acidosis in dairy cattle.

Dietary influence of de-oiled lecithin on broiler growth, Blood parameters, Antioxidant status, Thyroid hormones and immunoglobulins: An analysis across two distinct strains of broilers

The objective of this study was to examine the impact of adding an emulsifier, de-oiled soybean lecithin (DOL) in diet, on growth performance, blood parameters, antioxidant potential, thyroid hormones (T3 & T4), and serum immunoglobulins in two distinct strains of commercially raised broilers, Ross 308 (S1) and Evian 48 (S2). A total of 800 one-day-old broiler chicks from both strains (N = 400 each) were assigned to a control group (T0) and three dietary treatments, each with five replicates of 20 birds. Experimental treatments included a basal diet supplemented with varying levels of de-oiled lecithin (DOL): T1 = 0.5 g/kg, T2 = 1 g/kg, and T3 = 1.5 g/kg. The study used a 2 × 4 factorial design, which allows for the examination of the effects of two factors (in this case, strains and dietary treatments) on the outcome variables. Following 35 days, the broilers of strain S1 and those fed the T3 supplemented diet exhibited superior body weight (BW), body weight gain (BWG), cumulative feed intake (CFI), cumulative feed conversion ratio (CFCR), performance index (PI%), and lower mortality rate (Mr %) (P < 0.05) compared to those of strain S2 and those fed other dietary levels. A significant disparity (P < 0.05) was observed between the two strains regarding the plasma concentrations of ALB, GLB, A/G ratio, CHO, HDL, AST, and ALT, with strain S1 showing better levels. Similarly, the broiler chicks in group T3, along with the interaction between groups S1 × T3 and S2 × T3, demonstrated a significant (P < 0.05) improvement in blood biochemical parameters. Broilers of strain S1 and those on the T3 supplemented diet, along with the interactions between the groups S1 × T3 and S2 × T3, demonstrated substantial improvements (P < 0.05) across assessed parameters of antioxidant status and thyroid hormones, when compared to other dietary treatments and the control group. In conclusion, the addition of DOL as a supplement improved growth performance, blood parameters, antioxidant capacity, thyroid hormones (T3 & T4), and serum immunoglobulins in both broiler strains.

Genetic diversity assessment and selection of Bradyrhizobium strains for Inceptisols based on symbiotic performance

The symbiotic association of pigeonpea (Cajanus cajan (L.) Millsp.) with rhizobia plays a pivotal role in plants’ growth and development. The present study was carried out at Institute of Agricultural Sciences, Banaras Hindu University, Varanasi, Uttar Pradesh during 2018–2020 to explore the diversity of rhizobia present in the Cajanus cajan root under Inceptisols and identify an elite strain that exhibits exceptional plant growth promotion and of nitrogenase activity for efficient nitrogen fixation. The bacterial identification using 16s rDNA sequencing revealed bacteria strains Bradyrhizobium japonicum (S12), Bradyrhizobium subterraneum (S1, S7, S8, S10, S13), Bradyrhizobium yuanmingense (S2, S3, S4, S6, S9, S11) while two bacteria were the endophytes identified as Pseudomonas azotoformans (S5) and Paenibacillus seodonensis (S15). Genetic diversity using UPGMA clustering revealed remarkable genetic variations, classifying 14 strains into three primary clusters and four secondary clusters. Strains S1, S3, S6, S9, and S13 exhibited elevated nitrogenase activity, indicative of their proficient nitrogen fixation capability. In plant growth experiments, all strains demonstrated significant growth of the plants compared to control. The notable growth enhancement by these five strains can be attributed to their remarkable efficiency in nitrogen fixation, as indicated by significantly higher nodule formation and nitrogenase activity. These findings provide valuable insights for identifying potential rhizobial inoculants to enhance pigeonpea productivity.

Coat protein genealogy and complete genome characterization of field isolates of rice yellow mottle virus from Zambia

AbstractRice yellow mottle virus (RYMV) is widespread in mainland Africa and adjoining islands but to date its occurrence in Zambia is unknown. In March 2022, field surveys were conducted in Luapula, Northern, Western and Eastern provinces of Zambia to determine the occurrence of RYMV and its genetic relationship with global isolates of the virus. Thirty‐three paddy rice fields were visited and 108 leaf tissue samples were collected for analysis. The incidence of yellow mottle symptoms ranged from 25% to 43% in 10 (30.3%) of 33 fields and RYMV was detected in 35 (32.4%) of 108 samples by RT‐PCR with virus‐specific primers. A subset of 27 RYMV‐positive samples was constituted from which full‐length (720 bp) coat protein (CP) regions were amplified followed by bidirectional Sanger sequencing. Phylogenetic analysis of the CP cistron revealed distinct clustering of the isolates from Zambia in a monophyletic clade as subtype of the RYMV strain S4. Three isolates were randomly selected and used to obtain complete RYMV genomes of 4448–4449 nucleotides (nt). Comparative analysis of both the CP and the complete RYMV genomes from Zambia with their corresponding sequences in GenBank revealed that they shared 92%–93.2% nt identities with strain S4 isolates. These results confirm for the first time occurrence of RYMV strain S4 in Zambia and further reinforce the need for phytosanitary vigilance to safeguard rice production in Zambia.

Sugar Alcohol Sweetener Production by Yarrowia lipolytica Grown in Media Containing Glycerol.

Most of the world's annual production of mannitol is by chemical means, but, due to increasing demand for natural sweeteners, alternative production methods are being sought. The aim of the study was to screen Yarrowia lipolytica yeast strains and select culture conditions for the efficient and selective biosynthesis of mannitol from glycerol. From 21 strains examined in the shake-flask culture for mannitol biosynthesis from glycerol (100 g/L), three strains were selected-S2, S3, and S4-and further evaluated in batch bioreactor cultures with technical and raw glycerol (150 g/L). The best production parameters were observed for strain S3, which additionally was found to be the most resistant to NaCl concentration. Next, strain S3 was examined in batch culture with regard to the initial glycerol concentration (from 50 to 250 g/L). It was found that the substrate concentrations of 50 and 75 g/L resulted in the highest mannitol selectivity, about 70%. The fed-batch culture system proposed in this paper (performed in two variants in which glycerol was dosed in four portions of about 50 or 75 g/L) resulted in increased mannitol production, up to 78.5 g/L.

Statistical optimization of process variables for agarase production using Microbacterium sp. SS5 strain from non-marine sources

Agar oligosaccharides are thought to be valuable biomolecules with high bioactivity potential, along with a wide range of applications and advantages. The current study aimed to optimize the culture parameters required to produce agarase enzyme and agar oligosaccharides from industrial waste agar. Microbacterium spp. strain SS5 was isolated from a non-marine source and could synthesize oligo derivatives for use in a variety of industries ranging from food to pharmaceuticals. In addition, the strain and culture conditions were optimized to maximize extracellular agarase production. The bacterium grew best at pH 5.0 − 9.0, with an optimal pH of 7.5 − 8.0; temperatures ranging from 25 to 45 °C, with an optimal of 35 °C; and carbon and nitrogen concentrations of 0.5% each. Plackett-Burman experimental design and response surface methods were used to optimize various process parameters for agarase production by Microbacterium spp. strain SS5. Using the Plackett-Burman experimental design, eleven process factors were screened, and agar, beef extract, CaCl2, and beginning pH were found as the most significant independent variables affecting agarase production with confidence levels above 90%. To determine the optimal concentrations of the identified process factors on agarase production, the Box- Behnken design was used. Agarase production by Microbacterium spp. strain SS5 after optimization was 0.272 U/mL, which was determined to be greater than the result obtained from the basal medium (0.132 U/mL) before screening using Plackett-Burman and BBD with a fold increase of 2.06.

Pentaene macrolides AB023a and takanawaene C produced by Streptomyces xanthocidicus strain S3 for controlling pepper anthracnose

A natural product library consisting of the culture extracts of 814 actinomycete strains was screened for antifungal compounds that disrupt the cell integrity of plant pathogenic fungi using an adenylate kinase (AK) assay system. The culture extract of Streptomyces xanthocidicus strain S3 exhibited high AK activity against various plant pathogens. The active ingredients, AT-1 and AT-2, were isolated from the culture extract using a series of chromatographic procedures. Based on MS, UV, and NMR spectrometric analyses, the structures of AT-1 and AT-2 were determined as the pentaene macrolides, AB023a and takanawaene C. AB023a and takanawaene C displayed broad-spectrum antifungal activity against Aspergillus oryzae, Botrytis cinerea, Colletotrichum coccodes, C. gloeosporioides, C. orbiculare, Cylindrocarpon destructans, and Fusarium oxysporum f. sp. lycopersici, showing minimum inhibitory concentrations of 1–32 μg/mL. Treatment of AB023a and takanawaene C successfully inhibited anthracnose development on pepper plants in a concentration-dependent manner without phytotoxicity. The disease control efficacy of both compounds was comparable to that of the commercial fungicide chlorothalonil. Collectively, these results suggest that the polyene macrolides produced by S. xanthocidicus strain S3 can be used as natural fungicides for plant disease control.

Complete genome sequence of Roseinatronobacter sp. strain S2, a chemolithoheterotroph isolated from a pH 12 serpentinizing system.

Here, we report the complete genome sequence for Roseinatronobacter sp. S2, a sulfur-oxidizing heterotroph isolated from a serpentinizing system in Northern California. The S2 genome is 4.4 Mb and contains 4,570 protein-encoding genes. This organism contains the genes necessary for sulfur species oxidation and complete ethylmalonyl and pentose phosphate pathways.

Grain quality and yield response of rice to application of plant growth-promoting bacteria and amino acids

To investigate the effects of foliar application of plant growth-promoting bacteria (PGPB) and amino acids (AAs) on grain yield, nutrient concentrations and protein content of rice grain, a field experiment was conducted as a split-plot in a randomized complete block design with three replications during 2019-2020. The experimental treatments included the PGPB at five levels (control, P. agglomerans strain O4 (PGPB1), P. putida strain P13 + P. agglomerans strain P5 (PGPB2), P. koreensis strain S14 + P. vancouverensis strain S19 (PGPB3), and PGPB1 + PGPB2 + PGPB3 (PGPB4) as main plots and AAs at four levels (control, methionine, lysine, and methionine + lysine) as sub-plots. The results indicated that PGPB significantly enhanced the grain yield, protein content and nutrient uptake in rice grains when compared with control plants. However, the plants treated with co-application of PGPB strains (PGPB4) showed higher yield, higher protein accumulation and greater nutrients uptake such as N, P, K and Fe compared with individual application of PGPB strains and untreated plants. Among the separate application of PGPB strains, rice plants treated with PGPB1 with capability to N-fixing had the potential for enhance yield and improve protein and nutrients accumulation. The combined foliar application of AAs had greatest impact in increasing yield and improving rice grain quality in terms of protein content and uptake of nutrients, especially N, Ca and Fe in grain. Overall, our results suggest that co-applications of PGPB strains (PGPB4) and using a mixture of two AAs (methionine + lysine) may find value as plant growth-promoting for increasing rice yield and quality.

Isolation, characterization, and pathogenicity assessment of Corynebacterium pseudotuberculosis biovar equi strains from alpacas (Vicugna pacos) in China.

Corynebacterium pseudotuberculosis is a zoonotic pathogen that causes lymphadenitis in humans, livestock, and wildlife. In this study, C. pseudotuberculosis biovar equi strains were isolated from three alpacas. Antibiotic susceptibility tests and pathogenicity tests were also conducted. Moreover, one strain was sequenced using DNBSEQ and Oxford Nanopore technology. The three strains exhibited resistance to aztreonam, fosfomycin, and nitrofurantoin. The median lethal doses (LD50) of strains G1, S2 and BA3 in experimentally infected mice was 1.66 × 105 CFU, 3.78 × 105 CFU and 3.78 × 105 CFU, respectively. The sequencing of strain G1 resulted in the assembly of a chromosomal scaffold comprising 2,379,166 bp with a G + C content of 52.06%. Genome analysis of strain G1 revealed the presence of 48 virulence genes and 5 antibiotic resistance genes (ARGs). Comparative genomic analysis demonstrates a high degree of genetic similarity among C. pseudotuberculosis strains, in contrast to other Corynebacterium species, with a clear delineation between strains belonging to the two biovars (ovis and equi). The data of the present study contribute to a better understanding of the properties of C. pseudotuberculosis biovar equi strains and the potential risk they pose to alpacas and other livestock, as well as the necessity of ongoing surveillance and monitoring of infectious diseases in animals.

The advances of the Chinese Brucella suis strain 2 vaccine

AbstractBrucellosis, a worldwide crucial zoonotic disease, poses global socioeconomic risks to the system by threatening human health and animal reproduction. In recent years, with the increase of animal husbandry, animal brucellosis has become a remarkable health issue in China. To prevent and control the spread of brucellosis, Brucella suis live strain S2 vaccine has been licensed as a multivalent vaccine against the three major zoonotic Brucella species, for example, Brucella melitensis (small ruminants), Brucella abortus (bovids), and B. suis (swine). Following this process, the country enforced control measures based upon B. suis S2 vaccination, which reduced the disease burden in 2022 to 0.66% in the livestock population. The aim of this paper is to review and discuss the biological properties of the S2 vaccine and its wide use in vaccination programs of China, highlighting oral vaccination advantages, validation, and standardization of this vaccine as well as the progress of the S2 vaccination program achieved in the national control of the three zoonotic Brucella species.

Arsenic pollution and arsenic-resistant bacteria of drying Urmia Salt Lake

Extensive arsenic contamination of groundwater, rivers, and agricultural farms has led to widespread arsenic poisoning and consequent problems associated with health and economy. In recent years, many studies have been performed to understand bacterial arsenic detoxification and metabolism which has paved the way for bioremediation applications. This study attempted to isolate and characterize arsenic-resistant bacteria from the drying Urmia lake. As a result of the 16S rDNA analysis, six arsenic-resistant strains were identified as the members of Shouchella, Salipaludibacillus, and Evansella genera. For some of the strains, the maximum tolerance concentration for either arsenate or arsenite was considerably high, 320 and 16 mM, respectively. All the strains harbored the arsenate reductase gene (arsC). The arsenate permease (arsB) gene was identified in all strains except in strains S1, S12, and E15. The metabolic genes of respiratory arsenate reductase (arrB) and arsenite oxidase (arxA) were identified in none of these strains. This feature of the strains with the arsC gene region can be exploited to bioremediate arsenic from contaminated areas by using a two-step process. The second step can be carried out by the adsorption of arsenite to iron oxide or by precipitation with sulfide. Today, more and more ecosystems are being destroyed due to anthropogenic pollution, and it is important to reveal the negative effects and solutions of this situation on the ecosystems. So, the current study provides a potential source of bacteria for such studies that implement bioremediation practices to prevent arsenic catastrophe in vicinal territories, and reveals arsenic pollution in Urmia lake.

Characterization of a Haloalkaline Nocardiopsis sp. Strain S10 Isolated from Wastewater and Optimization of Culture Medium for Improving Production of Antifungal Compounds

Background: The emerging antibiotic resistant bacteria (ARB) and antibiotic resistance genes (ARGs) are a pressing global concern. Furthermore, the limited number of available antifungal agents requires the discovery of new antimicrobials. Introduction: This paper focused on the isolation and characterization of an actinobacterium designated S10, the study of its antimicrobial activity, on the one hand, and the optimization of antifungal production using statistical methods, on the other hand. Methods and Results: A haloalkaliphilic actinobacterium designated S10 was isolated from a wastewater treatment plant in the Souk-El-Tenine region (Bejaia, Algeria). This strain showed a broad spectrum of activity against bacteria (B. subtilis, L. innocua, M. luteus, S. aureus, MRSA, E. coli, P. aeruginosa, S. Typhi, and V. cholerae), molds (A. niger, G. geotricum, and F. oxysporum) and a pathogenic yeast (C. albicans). The extracellular enzymes producer Nocardiopsis sp. S10 was identified based on morphological, chemical, and 16S rRNA gene sequencing analysis. The best antimicrobial production was achieved on Mincer-Sea-Water (MSW) solid medium, and the antibiotics produced were extracted with ethyl acetate. For antifungal production, central composite design (CCD) and response surface methodology (RSM) were used to optimize the culture medium formulation. Conclusion: Optimal antifungal activity was achieved using a mixture of 2 g. L-1 starch, 2 g. L-1 of yeast extract and 2.75 g. L-1 of peptone at pH = 9.2. The application of the statistical approach stimulated a good yield of anti-C. albicans activity, corresponding to an increase of 72.22% compared to the non-optimized conditions.

Comparative genomics reveal distinct potential of Tamlana sp. S12 for algal polysaccharide degradation

IntroductionMacroalgae contain various polysaccharides that serve as nutrient sources Introduction: Macroalgae contain various polysaccharides that serve as nutrient sources for marine bacteria. Carbohydrate-active enzymes (CAZymes) are the primary feature of marine bacteria that utilize these polysaccharides. In this study, we describe Tamlana sp. S12, a novel strain of marine flavobacteria that can degrade alginate and Laminaria japonica biomass, isolated from the intestines of the sea cucumber Apostichopus japonicas collected at Weihai coast.MethodsWe sequenced the entire genome of strain S12 and constructed a phylogenetic tree using the core genome sequences of related strains. We determined the enzymatic activity of strain S12 using the DNS method and measured its growth curve under different carbon sources using spectrophotometry.ResultsStrain S12 degraded dehydrated L. japonica fragments as the sole nutrient source within 48h. Strain S12 harbors a diverse array of CAZymes at multiple polysaccharide utilization loci (PUL). One PUL encoding lyases from PL6, 7, and 17 families may be used for the degradation of alginate. Additionally, strain S12 harbors PULs encoding carrageenan- and agar-targeting CAZymes. Comparative analysis with related flavobacteria from Algibacter, Maribacter, and Zobellia showed shared CAZymes among these strains, potentially derived from a common ancestor and stably maintained within strains. Genomic signatures, algal degradation ability, and CAZyme patterns suggest that strain S12 has the potential to degrade complex algal polysaccharides.ConclusionThese results expand our knowledge of CAZymes and enrich our understanding of how marine Flavobacteriaceae adapt to marine algal polysaccharide environments. The availability of the genome of Tamlana sp. S12 will be beneficial for further analyses of marine Flavobacteriaceae.

Recombinant human adenovirus type 5 based vaccine candidates against GIIa- and GIIb-genotype porcine epidemic diarrhea virus induce robust humoral and cellular response in mice

Porcine epidemic diarrhea virus (PEDV) is a porcine enteropathogenic coronavirus causing severe watery diarrhea, vomiting, dehydration, and death in piglets. However, most commercial vaccines are developed based on the GI genotype strains, and have poor immune protection against the currently dominant GII genotype strains. Therefore, four novel replication-deficient human adenovirus 5-vectored vaccines expressing codon-optimized forms of the GIIa and GIIb strain spike and S1 glycoproteins were constructed, and their immunogenicity was evaluated in mice by intramuscular (IM) injection. All the recombinant adenoviruses generated robust immune responses, and the immunogenicity of recombinant adenoviruses against the GIIa strain was stronger than that of recombinant adenoviruses against the GIIb strain. Moreover, Ad-XT-tPA-Sopt-vaccinated mice elicited optimal immune effects. In contrast, mice immunized with Ad-XT-tPA-Sopt by oral gavage did not induce strong immune responses. Overall, IM administration of Ad-XT-tPA-Sopt is a promising strategy against PEDV, and this study provides useful information for developing viral vector-based vaccines.

Characterization and genome analysis of Acinetobacter oleivorans S4 as an efficient hydrocarbon-degrading and plant-growth-promoting rhizobacterium

Plant-growth-promoting rhizobacteria (PGPR) have received increasing attention for assisting phytoremediation. However, the effect of PGPR on total petroleum hydrocarbon (TPH) degradation and plant growth promotion and its underlying mechanism is not well understood. In this study, phenotypic analysis and whole genome sequencing were conducted to comprehensively characterize a newly isolated rhizobacterium strain S4, which was identified as Acinetobacter oleivorans, from a TPH-contaminated soil. The strain degraded 62.5% of initially spiked diesel (1%) in minimal media within six days and utilized n-alkanes with a wide range of chain length (i.e., C12 to C40). In addition, the strain showed phenotypic traits beneficial to plant growth, including siderophore production, indole-3-acetic acid synthesis and phosphate solubilization. Potential metabolic pathways and genes encoding proteins responsible for the phenotypic traits were identified. In a real TPH-contaminated soil, inoculation of Acinetobacter oleivorans S4 significantly enhanced the growth of tall fescue relative to the soil without inoculation. In contrast, inoculation of Bacillus sp. Z7, a hydrocarbon-degrading strain, showed a negligible effect on the growth of tall fescue. The removal efficiency of TPH with inoculation of Acinetobacter oleivorans S4 was significantly higher than those without inoculation or inoculation of Bacillus sp. Z7. These results suggested that traits of PGPR beneficial to plant growth are critical to assist phytoremediation. Furthermore, heavy metal resistance genes and benzoate and phenol degradation genes were found in the genome of Acinetobacter oleivorans S4, suggesting its application potential in broad scenarios.

Antibiotic Susceptibility Profiling of Human Pathogenic Staphylococcus aureus Strains Using Whole Genome Sequencing and Genome-Scale Annotation Approaches.

Staphylococcus species are major pathogens with increasing importance due to the rise in antibiotic resistance. Whole genome sequencing and genome-scale annotation are promising approaches to study the pathogenicity and dissemination of virulence factors in nosocomial methicillin-resistant and multidrug-resistant bacteria in intensive care units. Draft genome sequences of eight clinical S. aureus strains were assembled and annotated for the prediction of antimicrobial resistance genes, virulence factors, and phylogenetic analysis. Most of the studied S. aureus strains displayed multi-resistance toward the tested drugs, reaching more than seven drugs up to 12 in isolate S22. The mecA gene was detected in three isolates (S14, S21, and S23), mecC was identified in S8 and S9, and blaZ was commonly identified in all isolates except strain S23. Additionally, two complete mobile genomic islands coding for methicillin resistance SCCmec Iva (2B) were identified in strains S21 and S23. Numerous antimicrobial resistance genes (norA, norC, MgrA, tet(45), APH(3')-IIIa, and AAC(6')-APH(2″)) were identified in chromosomes of different strains. Plasmid analysis revealed the presence of blaZ, tetK, and ermC in different plasmid types, located in gene cassettes containing plasmid replicons (rep) and insertion sequences (IS). Additionally, the aminoglycoside-resistant determinants were identified in S1 (APH(3')-IIIa), while AAC(6)-APH(2″) was detected in strains S8 and S14. The trimethoprim (dfrC) resistance gene was detected in S. aureus S21, and the fosfomycin (fosB) resistance gene was detected only in S. aureus S14. We also noted that S. aureus S1 belongs to ST1-t127, which has been reported as one of the most frequent human pathogen types. Additionally, we noted the presence of rare plasmid-mediated mecC-MRSA in some of our isolates.

Characterization of a Riboflavin-Producing Mutant of Bacillus subtilis Isolated by Droplet-Based Microfluidics Screening.

Bacillus subtilis is one of the commonly used industrial strains for riboflavin production. High-throughput screening is useful in biotechnology, but there are still an insufficient number of articles focusing on improving the riboflavin production of B. subtilis by this powerful tool. With droplet-based microfluidics technology, single cells can be encapsulated in droplets. The screening can be carried out by detecting the fluorescence intensity of secreted riboflavin. Thus, an efficient and high-throughput screening method suitable for riboflavin production strain improvement could be established. In this study, droplet-based microfluidics screening was applied, and a more competitive riboflavin producer U3 was selected from the random mutation library of strain S1. The riboflavin production and biomass of U3 were higher than that of S1 in flask fermentation. In addition, the results of fed-batch fermentation showed that the riboflavin production of U3 was 24.3 g/L, an 18% increase compared with the parent strain S1 (20.6 g/L), and the yield (g riboflavin/100 g glucose) increased by 19%, from 7.3 (S1) to 8.7 (U3). Two mutations of U3 (sinRG89R and icdD28E) were identified through whole genome sequencing and comparison. Then they were introduced into BS168DR (parent of S1) for further analysis, which also caused riboflavin production to increase. This paper provides protocols for screening riboflavin-producing B. subtilis with droplet-based microfluidics technology and reveals mutations in riboflavin overproduction strains.