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Frontiers in Microbiology 2024A comprehensive strategy for microbial identification and contamination investigation during sterile drug manufacturing was innovatively established in this study,...
OBJECTIVE
A comprehensive strategy for microbial identification and contamination investigation during sterile drug manufacturing was innovatively established in this study, mainly based on MALDI-TOF MS for the identification and complemented by sequencing technology on strain typing.
METHODS
It was implemented to monitor the bacterial contamination of a sterile drug manufacturing facility, including its bacterial distribution features and patterns. In three months, two hundred ninety-two samples were collected covering multiple critical components of raw materials, personnel, environment, and production water.
RESULTS
Based on our strategy, the bacterial profile across the production process was determined: 241/292 bacterial identities were obtained, and spp. (40.25%), spp.(11.20%), spp. (8.30%), (5.81%), and spp. (4.56%) are shown to be the most dominant microbial contaminants. With 75.8% species-level and 95.4% genus-level identification capability, MALDI-TOF MS was promising to be a first-line tool for environmental monitoring routine. Furthermore, to determine the source of the most frequently occurring , which evidenced a widespread presence in the entire process, a more discriminating whole-genome SNP typing method was developed to track the transmission routes. Phylogenetic analysis based on SNP results indicated critical environment contamination is highly relevant to personnel flow in this case. The strain typing results provide robust and accurate information for the following risk assessment step and support effective preventive and corrective measures.
CONCLUSION
In general, the strategy presented in this research will facilitate the development of improved production and environmental control processes for the pharmaceutical industry, and give insights about how to provide more sound and reliable evidence for the optimization of its control program.
PubMed: 38410390
DOI: 10.3389/fmicb.2024.1327175 -
Microorganisms Feb 2024Since thermophilic microorganisms are valuable sources of thermostable enzymes, it is essential to recognize the potential toxicity of silver nanoparticles used in...
Since thermophilic microorganisms are valuable sources of thermostable enzymes, it is essential to recognize the potential toxicity of silver nanoparticles used in diverse industrial sectors. Thermophilic bacteria 2Cx, 3CA, 3CA1, FMB1, and FMB2-1 were selected, and their MIC and MBC values were assessed by treatment with AgNPs in a range of 62.5-1500 μg mL. The growth inhibition curves showed that the 2Cx, and FMB2-1 strains were more sensitive to AgNPs, demonstrating a reduction in population by 71.1% and 31.7% at 62.5 μg mL and by 82.9% and 72.8% at 250 μg mL, respectively. TEM and FT-IR analysis revealed that AgNPs caused structural damage, cytoplasmic leakage, and disruption of cellular integrity. Furthermore, cell viability showed a significant decrease alongside an increase in superoxide radical (SOR; O) production. β-galactosidase biosynthesis decreased to 28.8% level at 500 μg mL AgNPs for 2Cx, 32.2% at 250 μg mL for FMB1, and 38.8% only at 62.5 μg mL, but it was completely inhibited at 500 μg mL for 3CA. Moreover, FMB2-1 showed a significant decrease to 11.2% at 125 μg mL. This study is the first to reveal the toxic effects of AgNPs on thermophilic bacteria.
PubMed: 38399806
DOI: 10.3390/microorganisms12020402 -
Microorganisms Feb 2024Phosphorous (P) is widely used in agriculture; yet, P fertilizers are a nonrenewable resource. Thus, mechanisms to improve soil P bioavailability need to be found....
Phosphorous (P) is widely used in agriculture; yet, P fertilizers are a nonrenewable resource. Thus, mechanisms to improve soil P bioavailability need to be found. Legumes are efficient in P acquisition and, therefore, could be used to develop new technologies to improve soil P bioavailability. Here, we studied different species and varieties of legumes and their rhizosphere microbiome responses to low-P stress. Some varieties of common beans, cowpeas, and peas displayed a similar biomass with and without P fertilization. The rhizosphere microbiome of those varieties grown without P was composed of unique microbes displaying different levels of P solubilization and mineralization. When those varieties were amended with P, some of the microbes involved in P solubilization and mineralization decreased in abundance, but other microbes were insensitive to P fertilization. The microbes that decreased in abundance upon P fertilization belonged to groups that are commonly used as biofertilizers such as and The microbes that were not affected by P fertilization constitute unique species involved in P mineralization such as , and . These P-insensitive microbes could be used to optimize P utilization and drive future sustainable agricultural practices to reduce human dependency on a nonrenewable resource.
PubMed: 38399758
DOI: 10.3390/microorganisms12020353 -
Microorganisms Feb 2024Rich endophytic bacterial communities exist in fescue ( L.) and play an important role in fescue growth, cold tolerance, drought tolerance and antibiotic tolerance. To...
Rich endophytic bacterial communities exist in fescue ( L.) and play an important role in fescue growth, cold tolerance, drought tolerance and antibiotic tolerance. To screen for probiotics carried by fescue seeds, seven varieties were collected from three different regions of China for isolation by the milled seed method and analyzed for diversity and motility, biofilm and antibiotic resistance. A total of 91 bacterial isolates were obtained, and based on morphological characteristics, 36 representative dominant strains were selected for 16S rDNA sequencing analysis. The results showed that the 36 bacterial strains belonged to four phyla and nine genera. The Firmicutes was the dominant phylum, and , and were the dominant genera. Most of the strains had motility (80%) and were biofilm-forming (91.7%). In this study, 15 strains were capable of Indole-3-acetic acid (IAA) production, 24 strains were capable of nitrogen fixation, and some strains possessed amylase and protease activities, suggesting their potential for growth promotion. Determination of the minimum inhibitory concentration (MIC) against the bacteria showed that the strains were not resistant to tetracycline and oxytetracycline. (QY6, LH4, MS2) and (YY4) showed resistance to five antibiotics (ampicillin, kanamycin, erythromycin, sulfadiazine and rifampicin). Using Pearson correlation analysis, a significant correlation was found between motility and biofilm, and between biofilm and sulfadiazine. In this study, we screened two strains of (QY6, LH4) with excellent growth-promoting ability as well as broad-spectrum antibiotic resistance. which provided new perspectives for subsequent studies on the strong ecological adaptations of fescue, and mycorrhizal resources for endophytic bacteria and plant interactions.
PubMed: 38399732
DOI: 10.3390/microorganisms12020329 -
International Journal of Molecular... Feb 2024A potential strain, sp. JNUCC32, was isolated and subjected to whole-genome sequencing. Genome functional annotation revealed its active metabolic capabilities. This...
Isolation, Characterization, Genome Annotation, and Evaluation of Tyrosinase Inhibitory Activity in Secondary Metabolites of sp. JNUCC32: A Comprehensive Analysis through Molecular Docking and Molecular Dynamics Simulation.
A potential strain, sp. JNUCC32, was isolated and subjected to whole-genome sequencing. Genome functional annotation revealed its active metabolic capabilities. This study aimed to investigate the pivotal secondary metabolites in the biological system. Fermentation and extraction were performed, resulting in the isolation of seven known compounds: tryptophol (), 3-(4-hydroxyphenyl)propionic acid (), ferulic acid (), maculosin (), brevianamide F (), indole-3-acetic acid (), and butyric acid (). Tryptophol exhibited favorable pharmacokinetic properties and demonstrated certain tyrosinase inhibitory activity (IC = 999 μM). For further analysis of its inhibition mechanism through molecular docking and molecular dynamics (MD) simulation, tryptophol formed three hydrogen bonds and a pro-Michaelis complex with tyrosinase (binding energy = -5.3 kcal/mol). The MD simulation indicated favorable stability for the tryptophol-mushroom tyrosinase complex, primarily governed by hydrogen bond interactions. The crucial residues VAL-283 and HIS-263 in the docking were also validated. This study suggests tryptophol as a potential candidate for antibrowning agents and dermatological research.
Topics: Molecular Dynamics Simulation; Molecular Docking Simulation; Monophenol Monooxygenase; Enzyme Inhibitors; Alcohols; Indoles
PubMed: 38396889
DOI: 10.3390/ijms25042213 -
International Journal of Molecular... Feb 2024Screening of with antagonistic effects on paddy mold pathogens to provide strain resources for biological control of mold in L. screening of isolates antagonistic...
Screening of with antagonistic effects on paddy mold pathogens to provide strain resources for biological control of mold in L. screening of isolates antagonistic towards from rhizosphere soil of healthy paddy; classification and identification of antagonistic strains by biological characteristics and 16S rDNA sequence analysis; transcriptome sequencing after RNA extraction from Bacillus-treated ; and extraction of inhibitory crude proteins of by ammonium sulfate precipitation; inhibitory crude protein and spp. were treated separately for and observed by scanning electron microscopy (SEM). An antagonistic strain of , named B7, was identified as by 16S rDNA identification and phylogenetic evolutionary tree comparison analysis. Analysis of the transcriptome results showed that genes related to secondary metabolite biosynthesis such as antifungal protein were significantly downregulated. SEM results showed that the mycelium of underwent severe rupture after treatment with and antifungal proteins, respectively. In addition, the sporocarp changed less after treatment with , and the sporangium stalks had obvious folds. B7 has a good antagonistic effect against and has potential for biocontrol applications of paddy mold pathogens.
Topics: Paenibacillus polymyxa; Antifungal Agents; Phylogeny; Antibiosis; Bacillus; DNA, Ribosomal; Paenibacillus; Aspergillus
PubMed: 38396880
DOI: 10.3390/ijms25042195 -
Fish Physiology and Biochemistry Jun 2024The present study investigated the effect of replacing fishmeal (FM) with insect meal of Hermetia illucens (HI) in the diet of Sparus aurata farmed inshore on growth,...
The present study investigated the effect of replacing fishmeal (FM) with insect meal of Hermetia illucens (HI) in the diet of Sparus aurata farmed inshore on growth, gut health, and microbiota composition. Two isolipidic (18% as fed) and isoproteic (42% as fed) diets were tested at the farm scale: a control diet without HI meal and an experimental diet with 11% HI meal replacing FM. At the end of the 25-week feeding trial, final body weight, specific growth rate, feed conversion rate, and hepatosomatic index were not affected by the diet. Gross morphology of the gastrointestinal tract and the liver was unchanged and showed no obvious signs of inflammation. High-throughput sequencing of 16S rRNA gene amplicons (MiSeq platform, Illumina) used to characterize the gut microbial community profile showed that Proteobacteria, Fusobacteria, and Firmicutes were the dominant phyla of the gut microbiota of gilthead seabream, regardless of diet. Dietary inclusion of HI meal altered the gut microbiota by significantly decreasing the abundance of Cetobacterium and increasing the relative abundance of the Oceanobacillus and Paenibacillus genera. Our results clearly indicate that the inclusion of HI meal as an alternative animal protein source positively affects the gut microbiota of seabream by increasing the abundance of beneficial genera, thereby improving gut health and maintaining growth performance of S. aurata from coastal farms.
Topics: Animals; Sea Bream; Animal Feed; Gastrointestinal Microbiome; Diet; RNA, Ribosomal, 16S; Chenopodiaceae; Intestines; Animal Nutritional Physiological Phenomena
PubMed: 38386264
DOI: 10.1007/s10695-024-01314-9 -
The ISME Journal Jan 2024Microorganisms colonizing plant roots co-exist in complex, spatially structured multispecies biofilm communities. However, little is known about microbial interactions...
Microorganisms colonizing plant roots co-exist in complex, spatially structured multispecies biofilm communities. However, little is known about microbial interactions and the underlying spatial organization within biofilm communities established on plant roots. Here, a well-established four-species biofilm model (Stenotrophomonas rhizophila, Paenibacillus amylolyticus, Microbacterium oxydans, and Xanthomonas retroflexus, termed as SPMX) was applied to Arabidopsis roots to study the impact of multispecies biofilm on plant growth and the community spatial dynamics on the roots. SPMX co-culture notably promoted root development and plant biomass. Co-cultured SPMX increased root colonization and formed multispecies biofilms, structurally different from those formed by monocultures. By combining 16S rRNA gene amplicon sequencing and fluorescence in situ hybridization with confocal laser scanning microscopy, we found that the composition and spatial organization of the four-species biofilm significantly changed over time. Monoculture P. amylolyticus colonized plant roots poorly, but its population and root colonization were highly enhanced when residing in the four-species biofilm. Exclusion of P. amylolyticus from the community reduced overall biofilm production and root colonization of the three species, resulting in the loss of the plant growth-promoting effects. Combined with spatial analysis, this led to identification of P. amylolyticus as a keystone species. Our findings highlight that weak root colonizers may benefit from mutualistic interactions in complex communities and hereby become important keystone species impacting community spatial organization and function. This work expands the knowledge on spatial organization uncovering interspecific interactions in multispecies biofilm communities on plant roots, beneficial for harnessing microbial mutualism promoting plant growth.
Topics: In Situ Hybridization, Fluorescence; RNA, Ribosomal, 16S; Biofilms; Microbial Interactions; Symbiosis
PubMed: 38365935
DOI: 10.1093/ismejo/wrae012 -
Veterinary Medicine and Science Mar 2024The application of high throughput technologies has enabled unravelling of unique differences between healthy mares and mares with endometritis at transcriptomic and...
BACKGROUND
The application of high throughput technologies has enabled unravelling of unique differences between healthy mares and mares with endometritis at transcriptomic and proteomic levels. However, differences in the uterine microbiome are yet to be investigated.
OBJECTIVES
The present study was aimed at evaluating the differences in uterine microbiome between healthy mares and mares with endometritis.
METHODS
Low-volume lavage (LVL) samples were collected from the uterus of 30 mares classified into healthy (n = 15) and endometritis (n = 15) based on their reproductive history, intrauterine fluid accumulation, gross appearance of LVL samples, endometrial cytology and bacterial culture. The samples were subjected to 16S rRNA sequencing.
RESULTS
Notable differences in the uterine microbiome were observed between healthy mares and mares with endometritis at various taxonomic levels. In healthy mares, the most abundant phylum, class, order and family were Firmicutes, Bacilli, Bacillales and Paenibacillaceae, respectively. In contrast, the most abundant corresponding taxonomic levels in mares with endometritis were Proteobacteria, Gammaproteobacteria, Enterobacterales and Enterobacteriaceae, respectively. At the genus level, Brevibacillus and Paenibacillus were more abundant in healthy mares, whereas Escherichia, Salmonella and Klebsiella were more abundant in mares with endometritis. In healthy mares, Brevibacillus brevis was the most abundant species, followed by Brevibacillus choshinensis and Paenibacillus sp JDR-2. However, in mares with endometritis, Escherichia coli was the most abundant species, followed by Salmonella enterica and Klebsiella pneumoniae.
CONCLUSIONS
These results confirmed the previously reported presence of a uterine microbiome in healthy mares and helped unravel some alterations that occur in mares with endometritis. The findings can potentially help formulate new approaches to prevent or treat equine endometritis.
Topics: Horses; Animals; Female; Endometritis; Proteomics; RNA, Ribosomal, 16S; Uterus; Microbiota
PubMed: 38357732
DOI: 10.1002/vms3.1369 -
Synthetic and Systems Biotechnology Mar 2024Polymyxin B, produced by is used as the last line of defense clinically. In this study, exogenous mixture of precursor amino acids increased the level and proportion of...
Polymyxin B, produced by is used as the last line of defense clinically. In this study, exogenous mixture of precursor amino acids increased the level and proportion of polymyxin B1 in the total of polymyxin B analogs of CJX518-AC (PPAC) from 0.15 g/L and 61.8 % to 0.33 g/L and 79.9 %, respectively. The co-culture of strain PPAC and recombinant -leu01, which produces high levels of threonine, leucine, and isoleucine, increased polymyxin B1 production to 0.64 g/L. When strains PPAC and -leu01 simultaneously inoculated into an optimized medium with 20 g/L peptone, polymyxin B1 production was increased to 0.97 g/L. Furthermore, the polymyxin B1 production in the co-culture of strains PPAC and -leu01 increased to 2.21 g/L after optimized inoculation ratios and fermentation medium with 60 g/L peptone. This study provides a new strategy to improve polymyxin B1 production.
PubMed: 38348399
DOI: 10.1016/j.synbio.2024.01.015