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MBio Apr 2023When microbes grow in foreign nutritional environments, selection may enrich mutations in unexpected pathways connecting growth and homeostasis. An evolution experiment...
When microbes grow in foreign nutritional environments, selection may enrich mutations in unexpected pathways connecting growth and homeostasis. An evolution experiment designed to identify beneficial mutations in Burkholderia cenocepacia captured six independent nonsynonymous substitutions in the essential gene , which modifies tRNA by adding a lysine to the anticodon for faithful AUA recognition. Further, five additional mutants acquired mutations in tRNA, which strongly suggests that disrupting the TilS-tRNA interaction was subject to strong positive selection. Mutated TilS incurred greatly reduced enzymatic function but retained capacity for tRNA binding. However, both mutant sets outcompeted the wild type by decreasing the lag phase duration by ~3.5 h. We hypothesized that lysine demand could underlie fitness in the experimental conditions. As predicted, supplemental lysine complemented the ancestral fitness deficit, but so did the additions of several other amino acids. Mutant fitness advantages were also specific to rapid growth on galactose using oxidative overflow metabolism that generates redox imbalance, not resources favoring more balanced metabolism. Remarkably, 13 mutations also evolved in the long-term evolution experiment with Escherichia coli, including four fixed mutations. These results suggest that TilS or unknown binding partners contribute to improved growth under conditions of rapid sugar oxidation at the predicted expense of translational accuracy. There is growing evidence that the fundamental components of protein translation can play multiple roles in maintaining cellular homeostasis. Enzymes that interact with transfer RNAs not only ensure faithful decoding of the genetic code but also help signal the metabolic state by reacting to imbalances in essential building blocks like free amino acids and cofactors. Here, we present evidence of a secondary function for the essential enzyme TilS, whose only prior known function is to modify tRNA to ensure accurate translation. Multiple nonsynonymous substitutions in , as well as its cognate tRNA, were selected in evolution experiments favoring rapid, redox-imbalanced growth. These mutations alone decreased lag phase and created a competitive advantage, but at the expense of most primary enzyme function. These results imply that TilS interacts with other factors related to the timing of exponential growth and that tRNA-modifying enzymes may serve multiple roles in monitoring metabolic health.
Topics: Lysine; Amino Acyl-tRNA Synthetases; Pyrimidine Nucleosides; Bacteria; RNA, Transfer; Escherichia coli; Mutation; Amino Acids
PubMed: 36853041
DOI: 10.1128/mbio.00287-23 -
Biomedicines Feb 2023The characteristics and roles of microbes in the occurrence and development of pulmonary nodules are still unclear.
BACKGROUND
The characteristics and roles of microbes in the occurrence and development of pulmonary nodules are still unclear.
METHODS
We retrospectively analyzed the microbial mNGS results of BALF from 229 patients with pulmonary nodules before surgery, and performed a comparative analysis of lung flora between lung cancer and benign nodules according to postoperative pathology. The analysis also focused on investigating the characteristics of lung microbiota in lung adenocarcinomas with varying histopathology.
RESULTS
There were differences in lung microbiota between lung cancer and benign lung nodules. Bacterial diversity was lower in lung cancer than in benign lung nodules. Four species (, , and ) were enriched in lung cancer compared with the benign lung nodules. The areas under the ROC curves of these four species were all greater than 0.6, and the AUC of was 0.702, which had the highest diagnostic value for differentiating lung cancer from benign lung diseases. The significantly enriched microbiota varied with the different pathological subtypes of lung adenocarcinoma. , and displayed a trend of increasing from the benign lung disease group to the AIS group, MIA group and IAC group, whereas showed a downward trend.
CONCLUSION
Changes in the abundance of lung microbiota are closely related to the development of infiltrating adenocarcinoma. Our findings provide new insights into the relationship between the changes in lung microbiota and the development of lung cancer.
PubMed: 36831166
DOI: 10.3390/biomedicines11020631 -
MicrobiologyOpen Feb 2023Universal stress proteins (USPs) are ubiquitously expressed in bacteria, archaea, and eukaryotes and play a lead role in adaptation to environmental conditions. They...
Universal stress proteins (USPs) are ubiquitously expressed in bacteria, archaea, and eukaryotes and play a lead role in adaptation to environmental conditions. They enable adaptation of bacterial pathogens to the conditions encountered in the human niche, including hypoxia, oxidative stress, osmotic stress, nutrient deficiency, or acid stress, thereby facilitating colonization. We previously reported that all six USP proteins encoded within a low-oxygen activated (lxa) locus in Burkholderia cenocepacia showed increased abundance during chronic colonization of the cystic fibrosis (CF) lung. However, the role of USPs in chronic cystic fibrosis infection is not well understood. Structural modeling identified surface arginines on one lxa-encoded USP, USP76, which suggested it mediated interactions with heparan sulfate. Using mutants derived from the B. cenocepacia strain, K56-2, we show that USP76 is involved in host cell attachment. Pretreatment of lung epithelial cells with heparanase reduced the binding of the wild-type and complement strains but not the Δusp76 mutant strain, indicating that USP76 is directly or indirectly involved in receptor recognition on the surface of epithelial cells. We also show that USP76 is required for growth and survival in many conditions associated with the CF lung, including acidic conditions and oxidative stress. Moreover, USP76 also has a role in survival in macrophages isolated from people with CF. Overall, while further elucidation of the exact mechanism(s) is required, we can conclude that USP76, which is upregulated during chronic infection, is involved in bacterial survival within CF macrophages, a hallmark of Burkholderia infection.
Topics: Humans; Burkholderia cenocepacia; Cystic Fibrosis; Heat-Shock Proteins; Persistent Infection; Burkholderia Infections; Hypoxia
PubMed: 36825886
DOI: 10.1002/mbo3.1311 -
Pediatric Pulmonology May 2023Cepacia syndrome (CS) is an acute, necrotizing pneumonia with a high mortality rate, occurring in patients with cystic fibrosis (CF) infected with Burkholderia cepacia... (Review)
Review
BACKGROUND
Cepacia syndrome (CS) is an acute, necrotizing pneumonia with a high mortality rate, occurring in patients with cystic fibrosis (CF) infected with Burkholderia cepacia complex (BCC). Due to its low incidence, data on this condition are limited.
METHODS
We conducted a systematic review of the reported cases of CS by searching MEDLINE, Embase and the Cochrane Library to improve knowledge of this rare but potentially lethal condition.
RESULTS
We included 15 eligible articles, describing 18 cases (9 females) of CS. Median age at onset was 22 years (range: 10-60 years); median time to CS after first infection by BCC was 5 years (range: 1-26 years). Burkholderia cenocepacia was the most frequently reported causative agent. All patients received intravenous antibiotic treatment (most frequently including cotrimoxazole), while inhaled antibiotics were used in five patients (27.8%). Immunosuppressant agents were the most commonly prescribed supportive treatment (n = 7, 38.9%). Half of the patients died (9/18, 50%).
CONCLUSIONS
This study describes epidemiological, clinical characteristics, and prognosis of CS cases reported over the last 24 years. CS is a rare yet severe complication of BCC infection in patients with CF, which occurs several years after BCC colonization and has a negative outcome in 50% of the patients. Data are too scanty to identify the most effective therapeutic approach.
Topics: Female; Humans; Child; Adolescent; Young Adult; Adult; Middle Aged; Cystic Fibrosis; Anti-Bacterial Agents; Burkholderia cepacia complex; Prognosis; Trimethoprim, Sulfamethoxazole Drug Combination; Burkholderia Infections
PubMed: 36815622
DOI: 10.1002/ppul.26359 -
Nature Communications Feb 2023Expanding mitochondrial base editing tools with broad sequence compatibility is of high need for both research and therapeutic purposes. In this study, we identify a...
Expanding mitochondrial base editing tools with broad sequence compatibility is of high need for both research and therapeutic purposes. In this study, we identify a DddA homolog from Simiaoa sunii (Ddd_Ss) which can efficiently deaminate cytosine in DC context in double-stranded DNA (dsDNA). We successfully develop Ddd_Ss-derived cytosine base editors (DdCBE_Ss) and introduce mutations at multiple mitochondrial DNA (mtDNA) loci including disease-associated mtDNA mutations in previously inaccessible GC context. Finally, by introducing a single amino acid substitution from Ddd_Ss, we successfully improve the activity and sequence compatibility of DdCBE derived from DddA of Burkholderia cenocepacia (DdCBE_Bc). Our study expands mtDNA editing tool boxes and provides resources for further screening and engineering dsDNA base editors for biological and therapeutic applications.
Topics: Gene Editing; CRISPR-Cas Systems; Mitochondria; DNA, Mitochondrial; Cytosine
PubMed: 36797253
DOI: 10.1038/s41467-023-36600-2 -
Molecules (Basel, Switzerland) Feb 2023The inhibition of carbohydrate-lectin interactions is being explored as an efficient approach to anti adhesion therapy and biofilm destabilization, two alternative...
The inhibition of carbohydrate-lectin interactions is being explored as an efficient approach to anti adhesion therapy and biofilm destabilization, two alternative antimicrobial strategies that are being explored against resistant pathogens. BC2L-C is a new type of lectin from that binds (mammalian) fucosides at the -terminal domain and (bacterial) mannosides at the -terminal domain. This double carbohydrate specificity allows the lectin to crosslink host cells and bacterial cells. We have recently reported the design and generation of the first glycomimetic antagonists of BC2L-C, β-- or β--fucosides that target the fucose-specific terminal domain (BC2L-C-Nt). The low water solubility of the designed -fucosides prevented a full examination of this promising series of ligands. In this work, we describe the synthesis and biophysical evaluation of new L-fucosyl and L-galactosyl amides, designed to be water soluble and to interact with BC2L-C-Nt. The protein-ligand interaction was investigated by Saturation Transfer Difference NMR, Isothermal Titration Calorimetry and crystallographic studies. STD-NMR experiments showed that both fucosyl and galactosyl amides compete with α-methyl fucoside for lectin binding. A new hit compound was identified with good water solubility and an affinity for BC2L-C-Nt of 159 μM (ITC), which represents a one order of magnitude gain over α-methyl fucoside. The x-ray structure of its complex with BC2L-C-Nt was solved at 1.55 Å resolution.
Topics: Animals; Lectins; Burkholderia cenocepacia; Ligands; Amides; Fucose; Mammals
PubMed: 36771163
DOI: 10.3390/molecules28031494 -
Biotechnologia 2022Phytate-mineralizing bacteria (PMB) with plant growth-promoting activity can be considered as a potential biofertilizer for plant nutrition. PMB catalyzes the conversion...
Phytate-mineralizing bacteria (PMB) with plant growth-promoting activity can be considered as a potential biofertilizer for plant nutrition. PMB catalyzes the conversion of insoluble sugar phosphates, inositols, nucleic acids, phospholipids, nucleotides, phytate, and phytin into soluble forms that can be assimilated by plants. The present study aimed to isolate potential PMB from rhizospheric soils and to study their plant growth-promoting potential for the possible development of a potential phosphobacterium biofertilizer. For this purpose, 34 PMB isolates were isolated that showed potent phytate-mineralizing potential. These isolates were tested for their potential to solubilize tricalcium phosphate (TCP) and for various other plant growth-promoting activities. Significant differences were found among the isolates with regard to phytate mineralization and other plant growth-promoting characteristics. The bacterial isolates biochemically identified as , , , and exhibited high/medium P solubilization, medium/high phytohormone production, and medium/low siderophore and ammonia production. Among all these isolates, isolate A14 ( strain FDAARGOS_7) was the promising isolate with high TCP solubilization, medium phytate mineralization, high enzyme production, medium/high phytohormone production, and medium ammonia production. This strain also showed nitrogen fixation activity, zinc solubilizing potential, potassium solubilization, ACC deaminase production, and catalase production. Hence, it can be concluded that can be the potential candidate for biofertilizer development. Future studies are planned for exploring the role of PMB in biofertilizer formulations.
PubMed: 36606074
DOI: 10.5114/bta.2022.116204 -
BMC Microbiology Jan 2023Burkholderia cenocepacia is an opportunistic pathogen that can cause acute and chronic infections in patients with weakened immune systems and in patients with cystic...
BACKGROUND
Burkholderia cenocepacia is an opportunistic pathogen that can cause acute and chronic infections in patients with weakened immune systems and in patients with cystic fibrosis. B. cenocepacia is resistant to many antibiotics making treatment challenging. Consequently, there is a critical need for alternative strategies to treat B. cenocepacia infections such as using bacteriophages and/or bacteriophages with subinhibitory doses of antibiotic called phage-antibiotic synergy.
RESULTS
We isolated a bacteriophage, KP1, from raw sewage that infects B. cenocepacia. Its morphological characteristics indicate it belongs in the family Siphoviridae, it has a 52 Kb ds DNA genome, and it has a narrow host range. We determined it rescued infections in Lemna minor (duckweed) and moderately reduced bacterial populations in our artificial sputum medium model.
CONCLUSION
These results suggest that KP1 phage alone in the duckweed model or in combination with antibiotics in the ASMDM model improves the efficacy of reducing B. cenocepacia populations.
Topics: Humans; Burkholderia cenocepacia; Bacteriophages; Anti-Bacterial Agents; Burkholderia Infections
PubMed: 36600213
DOI: 10.1186/s12866-022-02738-0 -
Biofilm Dec 2022The biofilm state is the preferred lifestyle of bacteria in nature. Within a biofilm, the resident bacteria are protected from environmental stresses, antibiotics and...
The biofilm state is the preferred lifestyle of bacteria in nature. Within a biofilm, the resident bacteria are protected from environmental stresses, antibiotics and other antimicrobials, including those due to multiple immune effectors of their host during conditions of disease. Thereby, biofilms contribute significantly to pathogenicity, recalcitrance to clearance and chronicity/recurrence of bacterial diseases, including diseases of the respiratory tract. In the absence of highly effective, biofilm-targeted therapeutics, antibiotics are commonly prescribed to attempt to treat these diseases, however, in light of the canonical resistance of biofilm-resident bacteria to antibiotic-mediated killing, this ineffectual practice often fails to resolve the diseased condition and contributes significantly to the global threat of rising antimicrobial resistance. Nontypeable is a common respiratory tract disease co-pathogen, often present in partnership with other airway pathogens. Herein we aspired to determine whether either of two monoclonal antibodies we developed, one specific for NTHI [directed against the majority subunit (PilA) of the type IV pilus (T4P) of NTHI] and the other able to act agnostically on all bacteria tested to date (directed against a structural protein of the biofilm matrix, a DNABII protein), were able to disrupt 2-genera biofilms wherein NTHI co-partnered with another respiratory tract pathogen. These monoclonals were tested singly as well as when within an antibody cocktail. The monoclonal directed against the NTHI antigen PilA was only effective on single species NTHI biofilms and not on single species biofilms formed by other unrelated species. However, when NTHI co-partnered with any of 5 respiratory tract pathogens tested here (, ), this exclusively NTHI-directed monoclonal was able to disrupt these 2-genera biofilms. Conversely, the monoclonal antibody directed against protective epitopes of a DNABII protein, significantly disrupted all single species and 2-genera biofilms, which reflected the universal presence of this structural protein in all tested biofilm matrices. However, greatest release of both pathogens from a 2-genera biofilm was uniformly achieved by incubation with a 1:1 cocktail of both monoclonals. These data support the use of an approach wherein patients with respiratory tract disease could be treated with a therapeutic monoclonal antibody cocktail to release NTHI and its common co-pathogens from the protective biofilm to be killed by either traditional antibiotics and/or host immune effectors.
PubMed: 36532267
DOI: 10.1016/j.bioflm.2022.100096 -
Japanese Journal of Infectious Diseases Mar 2023Burkholderia pseudomallei, the highly infectious and causative organism of melioidosis, was first identified in Myanmar in 1911. B. pseudomallei was identified in...
Burkholderia pseudomallei, the highly infectious and causative organism of melioidosis, was first identified in Myanmar in 1911. B. pseudomallei was identified in Myanmar because of its genetic relatedness to Burkholderia species. In this study, we identified two isolates of Burkholderia cenocepacia, two Acinetobacter baumannii complexes, and 18 clinical isolates of B. pseudomallei using Vitek 2. These isolates were first screened using a latex agglutination test, which showed positive results in 20 of the 22 isolates. All isolates were cultured on Ashdown՚s agar and further tested using molecular methods. Specific PCR for type III secretion system (TTSs) gene clusters indicated 19 B. pseudomallei isolates out of 22 isolates. Furthermore, 16S rRNA and recA gene sequencing were used as the gold standard methods and yielded the same results. RapID NF Plus detected 16 B. pseudomallei out of 22 isolates. Vitek 2 and RapID NF Plus should be considered key tools in the diagnosis of melioidosis and surveillance of B. pseudomallei in Myanmar; however, accurate identification must be confirmed by TTS1 PCR. This study evaluated the presumptive workflow for the investigation of B. pseudomallei infections using different methods and options, in line with the available equipment.
Topics: Humans; Burkholderia pseudomallei; Melioidosis; RNA, Ribosomal, 16S; Myanmar; Workflow
PubMed: 36450576
DOI: 10.7883/yoken.JJID.2022.508