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1.
J Hazard Mater ; 433: 128711, 2022 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-35395524

RESUMO

The study aims to see how effective the Citrobacter species strain is in removing HgII under stressful conditions. For this, a response surface methodology was chosen to optimized pH, temperature, and biomass for effective biotransformation of HgII. The optimized value for pH, temperature, and biomass were 6.5, 30 °C, and 2 mg/l with 89% HgII removal potential. TEM-EDX showed accumulated mercury onto the bacterial surface. Pot study was conducted to check the potentiality of this strain in alleviating the toxicity in Solanum lycopersicum L. under different concentrations of mercury. The enhancement in antioxidative enzymes, as well as mercury accumulation, was observed in test plants inoculated with IITISM25. Obtained result showed a greater accumulation of mercury in the root system than that of the shoot system due to poor translocation. Moreover, mercury reductase enzyme synthesis was also boosted by the addition of ß-mercaptoethanol and L-cysteine. The optimized condition for maximum enzyme synthesis was at pH 7.5 and temperature 30 °C with Km = 48.07 µmol and Vmax = 9.75 µmol/min. Thus, we can say that Citrobacter species strain IITISM25 can be effectively applied in remediation of HgII stress condition as well as promotion of Solanum lycopersicum L growth under stress conditions as a promising host.


Assuntos
Citrobacter , Mercúrio , Antioxidantes/metabolismo , Biomassa , Biotransformação , Citrobacter/metabolismo , Mercúrio/metabolismo , Mercúrio/toxicidade
2.
Microbiol Spectr ; 10(2): e0150621, 2022 Apr 27.
Artigo em Inglês | MEDLINE | ID: mdl-35357225

RESUMO

During a surveillance study conducted to assess the occurrence and genomic landscape of critical priority pathogens circulating at the human-animal-environment interface in Brazil, as part of the Grand Challenges Explorations-New Approaches to Characterize the Global Burden of Antimicrobial Resistance program, two multidrug-resistant (MDR) Citrobacter portucalensis carrying blaCTX-M-15 extended-spectrum ß-lactamase (ESBL) genes, isolated from green sea turtles, were characterized. Genomic and phylogeographical analysis of C. portucalensis genomes available in public databases revealed the intercontinental dissemination of clades carrying different arrays of clinically relevant genes conferring resistance to carbapenems, broad-spectrum cephalosporins, cephamycins, aminoglycosides and fluoroquinolones, disinfectants, and heavy metals. Our observations suggest that C. portucalensis could be emerging as critical priority bacteria of both public and One Health importance worldwide. IMPORTANCE The global spread of antibiotic-resistant priority pathogens beyond the hospital setting is a critical issue within a One Health context that integrates the human-animal-environment interfaces. On the other hand, next-generation sequencing technologies along with user-friendly and high-quality bioinformatics tools have improved the identification of bacterial species, and bacterial resistance surveillance. The novel Citrobacter portucalensis species was proposed in 2017 after taxonomic reclassification and definition of the strain A60T isolated in 2008. Here, we presented genomic data showing the occurrence of multidrug-resistant C. portucalensis isolates carrying blaCTX-M-15 ESBL genes in South America. Additionally, we observed the intercontinental dissemination of clades harboring a broad resistome to clinically relevant antibiotics. Therefore, these findings highlight that C. portucalensis is a global MDR bacteria that carries intrinsic blaCMY- and qnrB-type genes and has become a critical priority pathogen due to the acquisition of clinically relevant resistance determinants, such as ESBL and carbapenemase-encoding genes.


Assuntos
Citrobacter , beta-Lactamases , Animais , Antibacterianos/farmacologia , Citrobacter/genética , Farmacorresistência Bacteriana Múltipla/genética , Testes de Sensibilidade Microbiana , beta-Lactamases/genética
3.
Biomed Res Int ; 2022: 6384742, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35309170

RESUMO

Whole-genome sequencing (WGS) data of a bacterial strain IITK SM2 isolated from an aquifer located in the middle Indo-Gangetic plain is reported here, along with its physiological, morphological, biochemical, and redox-transformation characteristics in the presence of dissolved arsenic (As). The aquifer exhibits oxidizing conditions relative to As speciation. Analyses based on 16S rRNA and recN sequences indicate that IITK SM2 was clustered with C. youngae NCTC 13708T and C. pasteuri NCTC UMH17T. However, WGS analyses using the digital DNA-DNA hybridization and Rapid Annotations using Subsystems Technology suggest that IITK SM2 belongs to a strain of C. youngae. This strain can effectively reduce As(V) to As(III) but cannot oxidize As(III) to As(V). It exhibited high resistance to As(V) [32,000 mg L-1] and As(III) [1,100 mg L-1], along with certain other heavy metals typically found in contaminated groundwater. WGS analysis also indicates the presence of As-metabolizing genes such as arsC, arsB, arsA, arsD, arsR, and arsH in this strain. Although these genes have been identified in several As(V)-reducers, the clustering of these genes in the forms of arsACBADR, arsCBRH, and an independent arsC gene has not been observed in any other Citrobacter species or other selected As(V)-reducing strains of Enterobacteriaceae family. Moreover, there were differences in the number of genes corresponding to membrane transporters, virulence and defense, motility, protein metabolism, phages, prophages, and transposable elements in IITK SM2 when compared to other strains. This genomic dataset will facilitate subsequent molecular and biochemical analyses of strain IITK SM2 to identify the reasons for high arsenic resistance in Citrobacter youngae and understand its role in As mobilization in middle Indo-Gangetic plain aquifers.


Assuntos
Arsênio , Água Subterrânea , Arsênio/análise , Citrobacter/genética , DNA , Água Subterrânea/química , RNA Ribossômico 16S/genética
4.
Biomed Res Int ; 2022: 5727638, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35155675

RESUMO

BACKGROUND: World Health Organization identified some Enterobacteriaceae as superbugs because of their high production and spread of extended-spectrum beta-lactamases (ESBL) and carbapenemases. Moreover, their resistance against commonly prescribed antibiotics left few choices of drugs to treat infection. This study is aimed at determining the magnitude of ESBL and carbapenemase-producing Enterobacteriaceae pathogens and their antimicrobial resistance pattern. MATERIALS AND METHODS: A hospital-based cross-sectional study was carried out from February to April 2019 in the Northwestern Ethiopia region. A total of 384 patients presumptive for bacterial infections were conveniently enrolled in the study. Specimens were collected and processed following standard bacteriological procedures. Drug susceptibility tests were performed using disk diffusion technique. ESBL and carbapenemase enzymes were tested by double disk diffusion and modified carbapenem inhibition methods, respectively. The data obtained were analyzed using SPSS version 22 software, and descriptive statistics were summarized in tables and graphs. RESULTS: Out of 384 clinical specimens processed 100 (26%) were culture positive for Enterobacteriaceae. The proportion of Enterobacteriaceae infection was relatively higher among in-patients 86 (32.6%) than out-patients 14 (11.7%). Overall, Escherichia coli 35 (9.1%) was the leading isolate followed by Klebsiella pneumoniae 31 (8.1%). Klebsiella pneumoniae 15 (15.6%) was the most frequent isolate from bloodstream infection and is the leading isolate from intensive care unit patients 15 (38.3%). Overall, 44 (44%) of Enterobacteriaceae were extended-spectrum beta-lactamase producers. Among them, Citrobacter spp. was the leading one 4 (80%) followed by Enterobacter cloacae 6 (60%) and K. pneumoniae 18 (58.1%). Furthermore, 6 (6%) of Enterobacteriaceae were carbapenemase-producers, in which 5 (50%) of E. cloacae and 3 (9.7%) of K. pneumoniae had highest percentage. Conclusions. ESBL and carbapenemase-producing isolates of Enterobacteriaceae are alarmingly spreading in the study area. Thus, improving the infection prevention strategy and further screening at the national level is recommended to develop the optimal use of antibiotics.


Assuntos
Carbapenêmicos/farmacologia , Farmacorresistência Bacteriana Múltipla , Infecções por Enterobacteriaceae/epidemiologia , Infecções por Enterobacteriaceae/microbiologia , Enterobacteriaceae/enzimologia , Enterobacteriaceae/isolamento & purificação , Proteínas de Bactérias/metabolismo , Enterobacteriáceas Resistentes a Carbapenêmicos/efeitos dos fármacos , Enterobacteriáceas Resistentes a Carbapenêmicos/isolamento & purificação , Citrobacter/efeitos dos fármacos , Citrobacter/isolamento & purificação , Estudos Transversais , Enterobacteriaceae/efeitos dos fármacos , Escherichia coli/efeitos dos fármacos , Escherichia coli/isolamento & purificação , Etiópia/epidemiologia , Humanos , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/isolamento & purificação , Testes de Sensibilidade Microbiana , Prevalência , beta-Lactamases/metabolismo
5.
Acta Crystallogr F Struct Biol Commun ; 78(Pt 2): 66-74, 2022 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-35102895

RESUMO

Hydrogenases catalyze the reversible oxidation of H2. Carbon monoxide (CO) is known to be a competitive inhibitor of O2-sensitive [NiFe]-hydrogenases. Although the activities of some O2-tolerant [NiFe]-hydrogenases are unaffected by CO, the partially O2-tolerant [NiFe]-hydrogenase from Citrobacter sp. S-77 (S77-HYB) is inhibited by CO. In this work, the CO-bound state of S77-HYB was characterized by activity assays, spectroscopic techniques and X-ray crystallography. Electron paramagnetic resonance spectroscopy showed a diamagnetic Ni2+ state, and Fourier-transform infrared spectroscopy revealed the stretching vibration of the exogenous CO ligand. The crystal structure determined at 1.77 Šresolution revealed that CO binds weakly to the nickel ion in the Ni-Fe active site of S77-HYB. These results suggest a positive correlation between O2 and CO tolerance in [NiFe]-hydrogenases.


Assuntos
Monóxido de Carbono/química , Citrobacter/enzimologia , Hidrogenase/antagonistas & inibidores , Hidrogenase/química , Proteínas de Bactérias/química , Monóxido de Carbono/metabolismo , Monóxido de Carbono/farmacologia , Domínio Catalítico , Cristalografia por Raios X , Espectroscopia de Ressonância de Spin Eletrônica , Inibidores Enzimáticos/química , Inibidores Enzimáticos/metabolismo , Inibidores Enzimáticos/farmacologia , Hidrogenase/metabolismo , Modelos Moleculares , Conformação Proteica , Espectroscopia de Infravermelho com Transformada de Fourier
6.
ACS Infect Dis ; 8(1): 59-65, 2022 01 14.
Artigo em Inglês | MEDLINE | ID: mdl-34979073

RESUMO

Non-antibiotic alternative treatments to combat the increasing number of infections caused by multidrug resistant bacteria are urgently needed. In recent years, bacteriophages have reemerged to potentially replace or complement the role of antibiotics, as bacterial viruses have the ability to inactivate pathogens. This study aimed to evaluate the synergy of phage-antibiotic combinations. A Citrobacter amalonaticus isolate was used in this study together with the phage MRM57. Eight different antibiotics with different mechanisms of action were used in combination with the phage to study the impact of the combination treatment on the minimal inhibitory concentrations. We found that antibiotic concentration dependent synergism exists, albeit at different extents, with very low numbers of phages. This demonstrates the use of phages as an adjuvant with a sublethal concentration of antibiotics as an effective therapeutic strategy.


Assuntos
Bacteriófagos , Antibacterianos/farmacologia , Citrobacter , Testes de Sensibilidade Microbiana
7.
Protein J ; 41(1): 131-140, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-35031980

RESUMO

Glucosinolates are plant natural products which on degradation by myrosinases give rise to the beneficial bioactive isothiocyanates. Recently, a myrosinase activity was detected in a Citrobacter strain isolated from soil. This enzyme was purified enabling its amino acid sequence and gene sequence (cmyr) to be determined. In order to study this myrosinase it was necessary to establish an expression system that would enable future work such as a structural determination of the protein to be carried out. The myrosinase gene was amplified, cloned and expressed in Escherichia coli with a 6XHis-tag. The heterologous expression of cmyr enabled relatively large amounts of myrosinase to be produced (3.4 mg cmyr/100 ml culture). Myrosinase activity was determined by mixing substrate and enzyme and determining glucose release. Optimum pH and temperature were determined to be pH 6.0 and 25 °C for the Ni-NTA purified protein. The kinetic parameters of the purified myrosinase were determined using sinigrin as a substrate. Km and Vmax were estimated as 0.18 mM and 0.033 mmol/min/mg respectively for sinigrin under optimum conditions and compared to other kinetic data for myrosinases. The substrate specificity of myrosinase was determined having the highest affinity for sinigrin followed by glucoiberin, progoitrin, glucoerucin, glucoraphanin and glucotropaeolin.


Assuntos
Citrobacter , Glucosinolatos , Citrobacter/genética , Citrobacter/metabolismo , Clonagem Molecular , Glucosinolatos/química , Glucosinolatos/metabolismo , Glicosídeo Hidrolases/química , Especificidade por Substrato
8.
Microb Drug Resist ; 28(2): 199-204, 2022 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-34520266

RESUMO

A carbapenem-resistant Citrobacter sedlakii strain AA2CS carrying blaNDM-5 was detected in outdoor aerosols of a wastewater treatment plant (WWTP) in China and the whole genome was sequenced subsequently. AA2CS was captured in an aerobic tank with aerosol particles of sizes ranging from 4.7 to 7.0 µm. Besides blaNDM-5, AA2CS also harbored 21 other antibiotic resistance genes and displayed a high level of resistance to ampicillin, cefotaxime, ceftazidime, tetracycline, and meropenem. BlaNDM-5 was located on the IncX3 plasmid (pCSNDM-5) with an IS3000-IS5-blaNDM-5-bleMBL-trpF-dsbD-IS26 structure. pCSNDM-5 was highly homologous to other blaNDM-5-carrying IncX3 plasmids in China and can be transferred to the Escherichia coli recipient J53. To our knowledge, this is the first report of carbapenem-resistant Enterobacteriaceae in outdoor aerosols in WWTPs.


Assuntos
Antibacterianos/farmacologia , Citrobacter/genética , Farmacorresistência Bacteriana Múltipla/genética , Genes Bacterianos/genética , Águas Residuárias/microbiologia , Aerossóis , Proteínas de Bactérias/genética , China , Testes de Sensibilidade Microbiana , Plasmídeos , beta-Lactamases/genética
9.
Sci Total Environ ; 806(Pt 3): 151336, 2022 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-34743821

RESUMO

A bacteria strain Citrobacter W4 isolated from the microalgae sewage culture system showed flocculating activity against Chlorella pyrenoidosa. In this work, operation parameters under outdoor conditions were optimized. When the bacterial-algal ratio was 4:1, G value was 26.30 s-1, and harvesting time was 6 h, the harvesting efficiency achieved 87.37 ± 2.96% without ions addition and pH adjustment. The microbial community structure analysis showed Citrobacter W4 was dominant in the harvesting process. Flocculating active substances were on the surface and metabolites of Citrobacter W4. The main component of bacteria flocculating active substances was protein. Polysaccharides and carboxylic acid also promoted flocculation. The flocculation mechanisms were mainly adsorption bridging, net catching, and sweeping, not electric neutralization. The quality of FAMEs was improved after flocculation. The cost of 1 kg dried microalgae flocculated by Citrobacter W4 was $1.35. The novel flocculating bacteria showed the potential to harvest microalgae cost-effectively and environmentally friendly.


Assuntos
Chlorella , Microalgas , Bactérias , Biomassa , Citrobacter , Floculação , Águas Residuárias
10.
Front Cell Infect Microbiol ; 11: 744431, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34858870

RESUMO

The rise of Carbapenem-resistant Enterobacterales (CRE) represents an increasing threat to patient safety and healthcare systems worldwide. Citrobacter spp., long considered not to be a classical nosocomial pathogen, in contrast to Klebsiella pneumoniae and Escherichia coli, is fast gaining importance as a clinical multidrug-resistant pathogen. We analyzed the genomes of 512 isolates of 21 CRE species obtained from 61 hospitals within a three-year-period and found that Citrobacter spp. (C. freundii, C. portucalensis, C. europaeus, C. koseri and C. braakii) were increasingly detected (n=56) within the study period. The carbapenemase-groups detected in Citrobacter spp. were KPC, OXA-48/-like and MBL (VIM, NDM) accounting for 42%, 31% and 27% respectively, which is comparable to those of K. pneumoniae in the same study. They accounted for 10%, 17% and 14% of all carbapenemase-producing CRE detected in 2017, 2018 and 2019, respectively. The carbapenemase genes were almost exclusively located on plasmids. The high genomic diversity of C. freundii is represented by 22 ST-types. KPC-2 was the predominantly detected carbapenemase (n=19) and was located in 95% of cases on a highly-conserved multiple-drug-resistance-gene-carrying pMLST15 IncN plasmid. KPC-3 was rarely detected and was confined to a clonal outbreak of C. freundii ST18. OXA-48 carbapenemases were located on plasmids of the IncL/M (pOXA-48) type. About 50% of VIM-1 was located on different IncN plasmids (pMLST7, pMLST5). These results underline the increasing importance of the Citrobacter species as emerging carriers of carbapenemases and therefore as potential disseminators of Carbapenem- and multidrug-resistance in the hospital setting.


Assuntos
Carbapenêmicos , Infecções por Enterobacteriaceae , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Carbapenêmicos/farmacologia , Citrobacter/genética , Infecções por Enterobacteriaceae/epidemiologia , Hospitais , Humanos , Klebsiella pneumoniae/genética , Testes de Sensibilidade Microbiana , Plasmídeos/genética , beta-Lactamases/genética
11.
Microbiol Spectr ; 9(3): e0163321, 2021 12 22.
Artigo em Inglês | MEDLINE | ID: mdl-34937176

RESUMO

The emergence of plasmid-mediated tigecycline resistance genes has attracted a great deal of attention globally. Currently, no comprehensive in-depth genomic epidemiology study of tet(X4)-bearing pathogens present of pork origin as the One Health approach has been performed. Herein, 139 fresh pork samples were collected from multiple regions in China and 58 tet(X4)-positive strains were identified. The tet(X4) gene mainly distributed in Escherichia coli (n = 55). Besides, 4 novel tet(X4)-positive bacterial species Klebsiella pneumoniae (n = 2), Klebsiella quasipneumoniae (n = 1), Citrobacter braakii (n = 1) and Citrobacter freundii (n = 1) were first characterized here. Four different core tet(X4)-bearing genetic environments and five types of tet(X4)-bearing tandem duplications were discovered among 58 strains. The results of the phylogenetic tree showed that there was some correlation between E. coli strains from pork, human, pig farms, and slaughterhouses. A total of seven types of plasmid replicons were found in tet(X4)-positive plasmids, among which multireplicon plasmids were observed. Notably, two tet(X4)-positive fusion plasmids pCSZ11R (IncX1-IncFIA-IncFIB-IncFIC) and pCSX5G-tetX4 (IncX1-IncFII-IncFIA) were formed by IS26 in the hot spot. Besides, six samples were identified to harbor two different tet(X4)-bearing strains. More interestingly, the absolute quantitative results showed that the expression levels of tet(X4) between different strains with different tet(X4) copies were approximate. In this study, the genetic environment of tet(X4)-positive plasmids containing different plasmid replicons was analyzed to provide a basis for the further development of effective control measures. It is also highlighted that animal-borne tet(X4)-bearing pathogens incur a transmission risk to consumed food. Therefore, there is an urgent need for large-scale monitoring as well as the development of effective control measures. IMPORTANCE Tigecycline was considered the last-line drug against serious infections caused by multidrug-resistant Gram-negative bacteria. However, the plasmid-mediated tigecycline resistance gene tet(X) has been widely reported in different sources of Enterobacterales and Acinetobacter in China. China is one of the largest pig-producing nations in the world, and in-depth investigation of gene in pork is vital to figure out the fundamental dissemination of these genes and set up a reasonable control framework. In this study, we conducted an in-depth and systematic analysis of the diversity of tet(X4)-positive plasmids and the genetic environment of tet(X4) contained in pork samples from multiple regions of China, providing a basis for further development of effective control measures. It is also highlighted that animal-borne tet(X4)-bearing pathogens incur a transmission risk to consumed food. Therefore, there is an urgent need for large-scale monitoring as well as the development of effective control measures.


Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana Múltipla/genética , Genes Bacterianos/genética , Genômica , Resistência a Tetraciclina/genética , Tigeciclina/farmacologia , Animais , Bactérias/genética , China , Citrobacter/genética , Citrobacter freundii/genética , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Escherichia coli/genética , Infecções por Escherichia coli/microbiologia , Proteínas de Escherichia coli/genética , Inocuidade dos Alimentos , Humanos , Klebsiella/genética , Klebsiella pneumoniae/genética , Testes de Sensibilidade Microbiana , Filogenia , Plasmídeos , Suínos , Tigeciclina/metabolismo
12.
mSphere ; 6(6): e0085021, 2021 12 22.
Artigo em Inglês | MEDLINE | ID: mdl-34730375

RESUMO

The spread of Klebsiella pneumoniae carbapenemase (KPC)-producing Enterobacterales is a public health concern. KPC-encoding blaKPC is predominantly spread by strains of a particular phylogenetic lineage, clonal group 258, but can also be spread by horizontal transfer of blaKPC-carrying plasmids. Here, we report the transfer of a blaKPC-2-harboring plasmid via mobilization from K. pneumoniae to Citrobacter freundii complex and Morganella morganii strains in a single patient. We performed draft whole-genome sequencing to analyze 20 carbapenemase-producing Enterobacterales strains (15 of K. pneumoniae, two of C. freundii complex, and three of M. morganii) and all K. pneumoniae strains using MiSeq and/or MinION isolated from a patient who was hospitalized in New York and Montreal before returning to Japan. All strains harbored blaKPC-2-containing Tn4401a. The 15 K. pneumoniae strains each belonged to sequence type 258 and harbored a Tn4401a-carrying multireplicon-type plasmid, IncN and IncR (IncN+R). Three of these K. pneumoniae strains also possessed a Tn4401a-carrying ColRNAI plasmid, suggesting that Tn4401a underwent interplasmid transposition. Of these three ColRNAI plasmids, two and one were identical to plasmids harbored by two Citrobacter europaeus and three M. morganii strains, respectively. The Tn4401a-carrying ColRNAI plasmids were each 23,753 bp long and incapable of conjugal transfer via their own genes alone, but they mobilized during the conjugal transfer of Tn4401a-carrying IncN+R plasmids in K. pneumoniae. Interplasmid transposition of Tn4401a from an IncN+R plasmid to a ColRNAI plasmid in K. pneumoniae and mobilization of Tn4401a-carrying ColRNAI plasmids contributed to the acquisition of blaKPC-2 in C. europaeus and M. morganii. IMPORTANCE Plasmid transfer plays an important role in the interspecies spread of carbapenemase genes, including the Klebsiella pneumoniae carbapenemase (KPC)-coding gene, blaKPC. We conducted whole-genome sequencing (WGS) analysis and transmission experiments to analyze blaKPC-2-carrying mobile genetic elements (MGEs) between the blaKPC-2-harboring K. pneumoniae, Citrobacter europaeus, and Morganella morganii strains isolated from a single patient. blaKPC-2 was contained within an MGE, Tn4401a. WGS of blaKPC-2-carrying K. pneumoniae, C. europaeus, and M. morganii strains isolated from one patient revealed that Tn4401a-carrying ColRNAI plasmids were generated by plasmid-to-plasmid transfer of Tn4401a from a multireplicon-type IncN and IncR (IncN+R) plasmid in K. pneumoniae strains. Tn4401a-carrying ColRNAI plasmids were incapable of conjugal transfer in C. europaeus and M. morganii but mobilized from K. pneumoniae to a recipient Escherichia coli strain during the conjugal transfer of Tn4401a-carrying IncN+R plasmid. Therefore, Tn4401a-carrying ColRNAI plasmids contributed to the acquisition of blaKPC-2 in C. europaeus and M. morganii.


Assuntos
Proteínas de Bactérias/genética , Citrobacter/genética , Transferência Genética Horizontal , Klebsiella pneumoniae/genética , Morganella morganii/genética , beta-Lactamases/genética , Citrobacter/enzimologia , Citrobacter/isolamento & purificação , Infecções por Enterobacteriaceae/microbiologia , Humanos , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/enzimologia , Klebsiella pneumoniae/isolamento & purificação , Testes de Sensibilidade Microbiana , Morganella morganii/enzimologia , Morganella morganii/isolamento & purificação , Plasmídeos , Sequenciamento Completo do Genoma
13.
mBio ; 12(5): e0241021, 2021 10 26.
Artigo em Inglês | MEDLINE | ID: mdl-34609899

RESUMO

The gut microbiota plays a crucial role in susceptibility to enteric pathogens, including Citrobacter rodentium, a model extracellular mouse pathogen that colonizes the colonic mucosa. C. rodentium infection outcomes vary between mouse strains, with C57BL/6 and C3H/HeN mice clearing and succumbing to the infection, respectively. Kanamycin (Kan) treatment at the peak of C57BL/6 mouse infection with Kan-resistant C. rodentium resulted in relocalization of the pathogen from the colonic mucosa and cecum to solely the cecal luminal contents; cessation of the Kan treatment resulted in rapid clearance of the pathogen. We now show that in C3H/HeN mice, following Kan-induced displacement of C. rodentium to the cecum, the pathogen stably colonizes the cecal lumens of 65% of the mice in the absence of continued antibiotic treatment, a phenomenon that we term antibiotic-induced bacterial commensalization (AIBC). AIBC C. rodentium was well tolerated by the host, which showed few signs of inflammation; passaged AIBC C. rodentium robustly infected naive C3H/HeN mice, suggesting that the AIBC state is transient and did not select for genetically avirulent C. rodentium mutants. Following withdrawal of antibiotic treatment, 35% of C3H/HeN mice were able to prevent C. rodentium commensalization in the gut lumen. These mice presented a bloom of a commensal species, Citrobacter amalonaticus, which inhibited the growth of C. rodentium in vitro in a contact-dependent manner and the luminal growth of AIBC C. rodentium in vivo. Overall, our data suggest that commensal species can confer colonization resistance to closely related pathogenic species. IMPORTANCE Gut bacterial infections involve three-way interactions between virulence factors, the host immune responses, and the microbiome. While the microbiome erects colonization resistance barriers, pathogens employ virulence factors to overcome them. Treating mice infected with kanamycin-resistant Citrobacter rodentium with kanamycin caused displacement of the pathogen from the colonic mucosa to the cecal lumen. Following withdrawal of the kanamycin treatment, 65% of the mice were persistently colonized by C. rodentium, which seemed to downregulate virulence factor expression. In this model of luminal gut colonization, 35% of mice were refractory to stable C. rodentium colonization, suggesting that their microbiotas were able to confer colonization resistance. We identify a commensal bacterium of the Citrobacter genus, C. amalonaticus, which inhibits C. rodentium growth in vitro and in vivo. These results show that the line separating commensal and pathogenic lifestyles is thin and multifactorial and that commensals may play a major role in combating enteric infection.


Assuntos
Citrobacter rodentium/crescimento & desenvolvimento , Citrobacter/fisiologia , Colo/microbiologia , Infecções por Enterobacteriaceae/microbiologia , Animais , Citrobacter rodentium/genética , Citrobacter rodentium/fisiologia , Feminino , Microbioma Gastrointestinal , Humanos , Mucosa Intestinal/microbiologia , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL
14.
Pak J Pharm Sci ; 34(3(Supplementary)): 1149-1156, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-34602445

RESUMO

As part of our continuous research to understand the interaction mechanism of drug and metallo-elements, heavy metal complexes of azithromycin (AZI) were synthesized with arsenic oxide, lead carbonate and silver chloride salts in molar ratio of 2: 1 (L: M). Synthesized heavy metal complexes have shown good percent yield and characterized through spectroscopic parameters including UV-Visible, TLC, FT-IR, NMR and elemental analysis (CHN). Spectroscopic characterization reveals the binding of ligand AZI with heavy metals in bi-dentate manner involving the hydroxide and 9a-NCH3 group of the aglycone ring of AZI. These newly synthesized heavy metal complexes were evaluated for their antimicrobial response against selected gram positive and gram negative organisms and antifungal species. It was noted that all newly synthesized complexes exhibits increased activity against B.subtilus whereas, AZI itself didn't show any activity, while synthesized complexes have low to moderate response against all the studied organisms. Complex A-M12 possess greater enzymatic response against both urease and alpha chymotrypsin among all the studied complexes. Results obtained were then statistically analyzed through one way ANOVA and Dunnett's test by using SPSS version 20.0 suggesting the significant response of complexes against selected organisms.


Assuntos
Antibacterianos/farmacologia , Antifúngicos/farmacologia , Trióxido de Arsênio/farmacologia , Azitromicina/farmacologia , Carbonatos/farmacologia , Complexos de Coordenação/farmacologia , Chumbo/farmacologia , Compostos de Prata/farmacologia , Trióxido de Arsênio/química , Azitromicina/análogos & derivados , Azitromicina/química , Bacillus subtilis/efeitos dos fármacos , Candida albicans/efeitos dos fármacos , Carbonatos/química , Quimotripsina/metabolismo , Citrobacter/efeitos dos fármacos , Complexos de Coordenação/química , Testes de Sensibilidade a Antimicrobianos por Disco-Difusão , Ensaios Enzimáticos , Escherichia coli/efeitos dos fármacos , Klebsiella pneumoniae/efeitos dos fármacos , Chumbo/química , Micrococcus luteus/efeitos dos fármacos , Proteus mirabilis/efeitos dos fármacos , Pseudomonas aeruginosa/efeitos dos fármacos , Salmonella typhi/efeitos dos fármacos , Shigella flexneri/efeitos dos fármacos , Compostos de Prata/química , Staphylococcus aureus/efeitos dos fármacos , Streptococcus/efeitos dos fármacos , Urease/metabolismo
15.
Front Cell Infect Microbiol ; 11: 737636, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34513738

RESUMO

Objectives: This prospective study was carried out to investigate molecular characteristics and antimicrobial susceptibility patterns of Citrobacter spp. from extraintestinal infections. Methods: Forty-six clinical Citrobacter spp. isolates were isolated from hospital patients with extraintestinal infections and analyzed by multilocus sequence typing (MLST) using seven housekeeping genes. Antimicrobial susceptibility testing was performed by disk diffusion method according to the Clinical and Laboratory Standards Institute (CLSI) recommendations. Adhesion and cytotoxicity to HEp-2 cells were assessed. Results: The 46 clinical Citrobacter spp. isolates were typed into 38 sequence types (STs), 9 of which belonged to four clonal complexes (CCs). None of the isolates shared the same ST or CCs with isolates from other countries or from other parts of China. Over half of the isolates were multidrug-resistant (MDR), with 17/26 C. freundii, 5/6 C. braakii, and 3/14 C. koseri isolates being MDR. Moreover, four isolates were carbapenem resistant with resistance to imipenem or meropenem. Among eight quinolone resistant C. freundii, all had a mutation in codon 59 (Thr59Ile) in quinolone resistance determining region of the gyrA gene. Only a small proportion of the isolates were found to be highly cytotoxic and adhesive with no correlation to sample sources. Conclusions: There was a diverse range of Citrobacter isolates causing extraintestinal infections and a high prevalence of MDR.


Assuntos
Antibacterianos , Citrobacter , Antibacterianos/farmacologia , Citrobacter/genética , Farmacorresistência Bacteriana/genética , Humanos , Testes de Sensibilidade Microbiana , Tipagem de Sequências Multilocus , Estudos Prospectivos , beta-Lactamases
16.
mBio ; 12(4): e0226521, 2021 08 31.
Artigo em Inglês | MEDLINE | ID: mdl-34465028

RESUMO

Phenazines are secreted metabolites that microbes use in diverse ways, from quorum sensing to antimicrobial warfare to energy conservation. Phenazines are able to contribute to these activities due to their redox activity. The physiological consequences of cellular phenazine reduction have been extensively studied, but the counterpart phenazine oxidation has been largely overlooked. Phenazine-1-carboxylic acid (PCA) is common in the environment and readily reduced by its producers. Here, we describe its anaerobic oxidation by Citrobacter portucalensis strain MBL, which was isolated from topsoil in Falmouth, MA, and which does not produce phenazines itself. This activity depends on the availability of a suitable terminal electron acceptor, specifically nitrate. When C. portucalensis MBL is provided reduced PCA and nitrate, it oxidizes the PCA at a rate that is environmentally relevant. We compared this terminal electron acceptor-dependent PCA-oxidizing activity of C. portucalensis MBL to that of several other gammaproteobacteria with various capacities to respire nitrate. We found that PCA oxidation by these strains in a nitrate-dependent manner is decoupled from growth and strain dependent. We infer that bacterial PCA oxidation is widespread and genetically determined. Notably, oxidizing PCA enhances the rate of nitrate reduction to nitrite by C. portucalensis MBL beyond the stoichiometric exchange of electrons from PCA to nitrate, which we attribute to C. portucalensis MBL's ability to also reduce oxidized PCA, thereby catalyzing a complete PCA redox cycle. This bidirectionality highlights the versatility of PCA as a biological redox agent. IMPORTANCE Phenazines are increasingly appreciated for their roles in structuring microbial communities. These tricyclic aromatic molecules have been found to regulate gene expression, be toxic, promote antibiotic tolerance, and promote survival under oxygen starvation. In all of these contexts, however, phenazines are studied as electron acceptors. Even if their utility arises primarily from being readily reduced, they need to be oxidized in order to be recycled. While oxygen and ferric iron can oxidize phenazines abiotically, biotic oxidation of phenazines has not been studied previously. We observed bacteria that readily oxidize phenazine-1-carboxylic acid (PCA) in a nitrate-dependent fashion, concomitantly increasing the rate of nitrate reduction to nitrite. Because nitrate is a prevalent terminal electron acceptor in diverse anoxic environments, including soils, and phenazine producers are widespread, this observation of linked phenazine and nitrogen redox cycling suggests an underappreciated role for redox-active secreted metabolites in the environment.


Assuntos
Proteínas de Bactérias/metabolismo , Citrobacter/metabolismo , Nitratos/metabolismo , Proteínas de Bactérias/genética , Biocatálise , Citrobacter/genética , Oxirredução , Fenazinas/metabolismo
17.
Recent Pat Biotechnol ; 15(4): 286-301, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34515017

RESUMO

BACKGROUND: L-Methioninase (EC 4.4.1.11; MGL) is a pyridoxal phosphate (PLP)-dependent enzyme that is produced by a variety of bacteria, fungi, and plants. L-methioninase, especially from Pseudomonas and Citrobacter sp., is considered as the efficient therapeutic enzyme, particularly in cancers such as glioblastomas, medulloblastoma, and neuroblastoma that are more sensitive to methionine starvation. OBJECTIVE: The low stability is one of the main drawbacks of the enzyme; in this regard, in the current study, different features of the enzyme, including phylogenetic, functional, and structural from Pseudomonas, Escherichia, Clostridium, and Citrobacter strains were evaluated to find the best bacterial L-Methioninase. METHODS: After the initial screening of L-Methioninase sequences from the above-mentioned bacterial strains, the three-dimensional structures of enzymes from Escherichia fergusonii, Pseudomonas fluorescens, and Clostridium homopropionicum were determined through homology modeling via GalaxyTBM server and refined by GalaxyRefine server. RESULTS AND CONCLUSION: Afterwards, PROCHECK, verify 3D, and ERRAT servers were used for verification of the obtained models. Moreover, antigenicity, allergenicity, and physico-chemical analysis of enzymes were also carried out. In order to get insight into the interaction of the enzyme with other proteins, the STRING server was used. The secondary structure of the enzyme is mainly composed of random coils and alpha-helices. However, these outcomes should further be validated by wet-lab investigations.


Assuntos
Proteínas de Bactérias/genética , Liases de Carbono-Enxofre/genética , Proteínas de Bactérias/química , Liases de Carbono-Enxofre/química , Citrobacter/enzimologia , Citrobacter/genética , Clostridium/enzimologia , Clostridium/genética , Escherichia/enzimologia , Escherichia/genética , Patentes como Assunto , Filogenia , Pseudomonas/enzimologia , Pseudomonas/genética
18.
J Biotechnol ; 339: 14-21, 2021 Sep 20.
Artigo em Inglês | MEDLINE | ID: mdl-34271055

RESUMO

Being able to recombine more than two genes with four or more crossover points in a sequence independent manner is still a challenge in protein engineering and limits our capabilities in tailoring enzymes for industrial applications. By computational analysis employing multiple sequence alignments and homology modeling, five fragments of six phytase genes (sequence identities 31-64 %) were identified and efficiently recombined through phosphorothioate-based cloning using the PTRec method. By combinatorial recombination, functional phytase chimeras containing fragments of up to four phytases were obtained. Two variants (PTRec 74 and PTRec 77) with up to 32 % improved residual activity (90 °C, 60 min) and retained specific activities of > 1100 U/mg were identified. Both variants are composed of fragments from the phytases of Citrobacter braakii, Hafnia alvei and Yersinia mollaretii. They exhibit sequence identities of ≤ 80 % to their parental enzymes, highlighting the great potential of DNA recombination strategies to generate new enzymes with low sequences identities that offer opportunities for property right claims.


Assuntos
6-Fitase , 6-Fitase/genética , Citrobacter/enzimologia , Estabilidade Enzimática , Hafnia alvei/enzimologia , Concentração de Íons de Hidrogênio , Proteínas Recombinantes de Fusão , Yersinia/enzimologia
19.
Curr Opin Microbiol ; 63: 76-82, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34243134

RESUMO

Enteric bacterial infections impose a significant and global health burden on society, and their threat is increasing in concert with a rise in antibiotic resistance. There is thus a great need to quickly develop new antimicrobial treatments and interest is growing in targeting pathogen nutrition and metabolism. In this review, we highlight recent research on the metabolism of Citrobacter rodentium, a murine-specific relative of enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic E. coli (EHEC). We focus on the mechanisms by which C. rodentium acquires nutrients as well as the distinct metabolic strategies that C. rodentium employs in varying spatiotemporal niches. We propose that identifying and targeting nutrients found essential for bacterial pathogenesis is an attractive anti-microbial approach in the new post-antibiotic era.


Assuntos
Infecções por Enterobacteriaceae , Escherichia coli Êntero-Hemorrágica , Escherichia coli Enteropatogênica , Animais , Citrobacter , Citrobacter rodentium , Refeições , Camundongos
20.
Antonie Van Leeuwenhoek ; 114(8): 1285-1292, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34117562

RESUMO

A novel arsenate-reducing bacterium, LY-1T, was isolated from freshwater sediment in Huangshi, China. Morphological analysis indicated that the cells were shaped like rods and were gram-negative. The major fatty acids (> 10%) were C16:0, summed feature 3 (C16:1 ω7c, C16:1 ω6c) and summed feature 8 (C18:1 ω7c, C18:1 ω6c). An assessment of the phylogeny based on 16S rRNA gene sequences indicated that the strain LY-1T belonged to the genus Citrobacter, while further analysis based on the recN gene indicated that LY-1T occupies a distinct phylogenetic niche within the Citrobacter genus. Moreover, average nucleotide identity and digital DNA-DNA hybridization between the strain LY-1T and the type strains of closely related species of the genus Citrobacter (C. europaeus, C. brakii, C. portucalensis, C. freundii, C. werkmanii, C. cronae, C. youngae, C. pasteurii, C. tructae, C. gillenii, and C. murliniae) were 85.8-93.8% and 31.2-56.9%, respectively. In addition, the LY-1T strain's capacity to metabolize various compounds and its characteristic G + C content of 51.9% were also distinct from other species of the Citrobacter genus. These discriminatory features cumulatively indicate the LY-1T strain as a new species within the Citrobacter genus. We propose the species name Citrobacter arsenatis for this new species, with LY-1T (= CCTCC AB 2019169T = KCTC 72440T) as the type strain.


Assuntos
Arseniatos , Citrobacter , Técnicas de Tipagem Bacteriana , Composição de Bases , Citrobacter/genética , DNA Bacteriano/genética , Ácidos Graxos/análise , Água Doce , Hibridização de Ácido Nucleico , Fosfolipídeos , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA
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