The Integrative and Conjugative Element ICECspPOL2 Contributes to the Outbreak of Multi-Antibiotic-Resistant Bacteria for Chryseobacterium Spp. and Elizabethkingia Spp.

Antibiotic resistance genes (ARGs) and horizontal transfer of ARGs among bacterial species in the environment can have serious clinical implications as such transfers can lead to disease outbreaks from multidrug-resistant (MDR) bacteria. Infections due to antibiotic-resistant Chryseobacterium and Elizabethkingia in intensive care units have been increasing in recent years. In this study, the multi-antibiotic-resistant strain Chryseobacterium sp. POL2 was isolated from the wastewater of a livestock farm. Whole-genome sequencing and annotation revealed that the POL2 genome encodes dozens of ARGs. The integrative and conjugative element (ICE) ICECspPOL2, which encodes ARGs associated with four types of antibiotics, including carbapenem, was identified in the POL2 genome, and phylogenetic affiliation analysis suggested that ICECspPOL2 evolved from related ICEEas of Elizabethkingia spp. Conjugation assays verified that ICECspPOL2 can horizontally transfer to Elizabethkingia species, suggesting that ICECspPOL2 contributes to the dissemination of multiple ARGs among Chryseobacterium spp. and Elizabethkingia spp. Because Elizabethkingia spp. is associated with clinically significant infections and high mortality, there would be challenges to clinical treatment if these bacteria acquire ICECspPOL2 with its multiple ARGs, especially the carbapenem resistance gene. Therefore, the results of this study support the need for monitoring the dissemination of this type of ICE in Chryseobacterium and Elizabethkingia strains to prevent further outbreaks of MDR bacteria. IMPORTANCE Infections with multiple antibiotic-resistant Chryseobacterium and Elizabethkingia in intensive care units have been increasing in recent years. In this study, the mobile integrative and conjugative element ICECspPOL2, which was associated with the transmission of a carbapenem resistance gene, was identified in the genome of the multi-antibiotic-resistant strain Chryseobacterium sp. POL2. ICECspPOL2 is closely related to the ICEEas from Elizabethkingia species, and ICECspPOL2 can horizontally transfer to Elizabethkingia species with the tRNA-Glu-TTC gene as the insertion site. Because Elizabethkingia species are associated with clinically significant infections and high mortality, the ability of ICECspPOL2 to transfer carbapenem resistance from environmental strains of Chryseobacterium to Elizabethkingia is of clinical concern.

Comparison of three species of Elizabethkingia genus by whole-genome sequence analysis.

Elizabethkingia are found to cause severe neonatal meningitis, nosocomial pneumonia, endocarditis and bacteremia. However, there are few studies on Elizabethkingia genus by comparative genomic analysis. In this study, three species of Elizabethkingia were found: E. meningoseptica, E. anophelis and E. miricola. Resistance genes and associated proteins of seven classes of antibiotics including beta-lactams, aminoglycosides, macrolides, tetracyclines, quinolones, sulfonamides and glycopeptides, as well as multidrug resistance efflux pumps were identified from 20 clinical isolates of Elizabethkingia by whole-genome sequence. Genotype and phenotype displayed a good consistency in beta-lactams, aminoglycosides and glycopeptides, while contradictions exhibited in tetracyclines, quinolones and sulfonamides. Virulence factors and associated genes such as hsp60 (htpB), exopolysaccharide (EPS) (galE/pgi), Mg2+ transport (mgtB/mgtE) and catalase (katA/katG) existed in all clinical and reference strains. The functional analysis of the clusters of orthologous groups indicated that 'metabolism' occupied the largest part in core genome, 'information storage and processing' was the largest group in both accessory genome and unique genome. Abundant mobile elements were identified in E. meningoseptica and E. anophelis. The most significant finding in our study was that a single clone of E. anophelis had been circulating within diversities of departments in a clinical setting for nearly 18 months.

Chryseobacterium gleum Isolation from Respiratory Culture Following Community-Acquired Pneumonia.

BACKGROUND Chryseobacterium gleum (C. gleum) is a rare but concerning device-associated infection that can cause urinary tract infections and pneumonia. It produces a biofilm and has intrinsic resistance to a wide array of broad-spectrum agents. Risk factors include neonate or immunocompromised states, intensive care unit admission for more than 21 days, broad-spectrum antibiotic exposure, indwelling devices, and mechanical ventilation. CASE REPORT A 61-year-old cachectic man presented in the United States with community-acquired pneumonia and immediately decompensated, requiring ventilator support. Despite starting broad-spectrum antibiotics, the patient developed fever, leukocytosis, and additional desaturation episodes. The patient's respiratory culture grew numerous C. gleum and few Stenotrophomonas (Xanthomonas) maltophilia. He also had a positive urine streptococcal pneumonia antigen. Broad-spectrum agents were discontinued after prolonged treatment due to a continued worsening clinical picture, and the patient was started on trimethoprim-sulfamethoxazole to cover C. gleum. The patient showed rapid clinical improvement on trimethoprim-sulfamethoxazole, with resolution of symptoms on post-discharge follow-up. CONCLUSIONS To the best of our knowledge, this is the first case report of a documented case of a patient with C. gleum respiratory infection successfully treated solely with trimethoprim-sulfamethoxazole. The expedient identification of C. gleum is essential for proper treatment. The literature has consistently shown isolated respiratory C. gleum strains to be largely susceptible to fluoroquinolones, piperacillin-tazobactam, or trimethoprim-sulfamethoxazole.

Chryseobacterium/Elizabethkingia species infections in Saudi Arabia.

OBJECTIVES: To describe the epidemiological, clinical, and outcome data of patients infected or colonized with Chryseobacterium/Elizabethkingia spp including antibiotic susceptibility patterns. METHODS: This retrospective study was conducted at Prince Sultan Military Medical City, Riyadh,  Saudi Arabia. All patients infected or colonized by Chryseobacterium /Elizabethkingia spp who were admitted between June 2013 and May 2019 were included. Data were extracted from patient electronic medical records. RESULTS: We enrolled 27 patients (13 males and 14 females) with a mean age of 35.6 years. Chryseobacterium/Elizabethkingia spp were isolated from blood cultures (n=13, 48%) and tracheal aspirations (n=11, 41%). The most frequent species isolated was Elizabethkingia meningoseptica (n=22). Although 6 patients were considered colonized, the remaining 21 patients presented with ventilator associated pneumonia (n=9), central line associated bloodstream infection (n=4), septic shock (n=4), or isolated bacteremia (n=4). In 25 cases the infections were health-care related. Three patients (11%) died within 28 days. Twenty-six isolates (96.5%) were resistant to carbapenems. Moxifloxacin and cotrimoxazole were the most active antibiotics. CONCLUSION: Chryseobacterium/Elizabethkingia spp infection is rare, but can be responsible for severe hospital acquired infections. Cotrimoxazole and fluoroquinolone are the most effective antibiotic treatments.

The regulatory mechanism of Chryseobacterium sp. resistance mediated by montmorillonite upon cadmium stress.

Cadmium (Cd) is a toxic heavy metal and its uptake by living organisms causes adverse effect, further resulting in cycle pollution of the biosphere. The specific regulatory mechanism between clays and microbes under Cd stress remains unclear. In this study, interface interactions among clays, microbes and Cd were confirmed. Comparative transcriptome was conducted to investigate how it regulated gene expression patterns of microbes (Chryseobacterium sp. WAL2), which exposed to a series of gradient concentrations of Cd (16, 32, 64 and 128 µg mL-1) for 12 d in the presence and absence of clay montmorillonite (Mt) (16 g L-1). Cd was highly enriched by the unique interface interactions between Mt and bacteria (67.6-82.1%), leading to a more hostile environment for bacterial cells. However, Mt ultimately enhanced bacterial resistance to Cd stress by stimulating the mechanism of bacterial resistance; namely: (i) Mt increased genes expression connected with ion transport, enhancing the uptake of Cd; (ii) Mt stimulated genes expression related to efflux pump and positively regulated cellular oxidative stress (e.g., glutathione) and Cd accumulation (e.g., cysteine) processes. Further, genes expression related to intracellular metabolic processes was enforced, which supplied a driving force and accelerated electron transfer; (iii) Mt improved genes expression involved in DNA replication and other biological processes (e.g., terpenoid backbone biosynthesis) to maintain bacterial vitality. Therefore, the study not only optimized a unique Cd resistance mechanism of Mt on Chryseobacterium sp., but also provided a novel insight for environmental mitigation of heavy metals from the perspective of molecular biology.

Microbiology of Elizabethkingia spp. isolates in hospitalized patients.

An increasing number of infections due to Elisabethkingia spp. have been observed and reported in recent years. Here we report the microbiological aspects of 13 cases with positive cultures for Elizabethkingia spp. from 1998 to 2017. Elizabethkingia isolates were identified using the Vitek 2 Compact 60 (AES software) Gram-Negative Identification test (GNI) card (bioMerieux. Marcy-l'Etoile, France). The MICs were determined using Vitek 2 and interpreted according to CLSI guidelines based on the interpretive MIC breakpoints for Acinetobacter spp. Vitek 2 susceptibility results were confirmed using the manual E-test and the colistin result was confirmed using the broth dilution method. Twelve cases were observed between 2010 and 2017 with four cases in 2017. Of the 2017 samples, three out of four isolates were obtained outside the intensive care units compared to one out of eight samples between 2010 and 2016. There was variable susceptibility to trimethoprim/sulfamethoxazole (58.3%), ciprofloxacin (41.7%), piperacillin/tazobactam and gentamicin (16.6% each), and all were resistant to colistin. There is a high rate of drug resistance and further studies to identify the source of the infection are needed to build up a profile of Elizabethkingia spp. to inform public health policy in this context.

Global Proteome Profiling Reveals Drug-Resistant Traits in Elizabethkingia meningoseptica: An Opportunistic Nosocomial Pathogen.

Elizabethkingia meningoseptica is Gram-negative, rod-shaped opportunistic bacterial pathogen increasingly reported in hospital-acquired outbreaks. This bacterium is well known to thrive in the hospital environment. One of the leading causes of meningitis in pediatric and immune-compromised patients, E. meningoseptica has been noted as a "pathogen of interest" in the context of nosocomial diseases associated with device-related infections in particular. This pathogen's multidrug-resistant phenotype and attendant lack of adequate molecular mechanistic data limit the current approaches for its effective management in hospitals and public health settings. This study provides the global proteome of E. meningoseptica. The reference strain E. meningoseptica ATCC 13253 was used for proteomic analysis using high-resolution Fourier transform mass spectrometry. The study provided translational evidence for 2506 proteins of E. meningoseptica. We identified multiple metallo-ß-lactamases, transcriptional regulators, and efflux transporter proteins associated with multidrug resistance. A protein Car D, which is an enzyme of the carbapenem synthesis pathway, was also discovered in E. meningoseptica. Further, the proteomics data were harnessed for refining the genome annotation. We discovered 39 novel protein-coding genes and corrected four existing translations using proteogenomic workflow. Novel translations reported in this study enhance the molecular data on this organism, thus improving current databases. We believe that the in-depth proteomic data presented in this study offer a platform for accelerated research on this pathogen. The identification of multiple proteins, particularly those involved in drug resistance, offers new future opportunities to design novel and specific antibiotics against infections caused by E. meningoseptica.

Genomic Features, Comparative Genomic Analysis, and Antimicrobial Susceptibility Patterns of Chryseobacterium arthrosphaerae Strain ED882-96 Isolated in Taiwan.

Bacteria belonging to the genus Chryseobacterium are ubiquitously distributed in natural environments, plants, and animals. Except C. indologenes and C. gleum, other Chryseobacterium species rarely cause human diseases. This study reported the whole-genome features, comparative genomic analysis, and antimicrobial susceptibility patterns of C. arthrosphaerae ED882-96 isolated in Taiwan. Strain ED882-96 was collected from the blood of a patient who had alcoholic liver cirrhosis and was an intravenous drug abuser. This isolate was initially identified as C. indologenes by using matrix-assisted laser desorption ionization-time of flight mass spectrometry. The analysis of 16S ribosomal RNA gene sequence revealed that ED882-96 shared 100% sequence identity with C. arthrosphaerae type strain CC-VM-7T. The results of whole-genome sequencing of ED882-96 showed two chromosome contigs and one plasmid. The total lengths of the draft genomes of chromosome and plasmid were 4,249,864 bp and 435,667 bp, respectively. The findings of both in silico DNA-DNA hybridization and average nucleotide identity analyses clearly demonstrated that strain ED882-96 was a species of C. arthrosphaerae. A total of 83 potential virulence factor homologs were predicted in the whole-genome sequencing of strain ED882-96. This isolate was resistant to all tested antibiotics, including ß-lactams, ß-lactam/ß-lactamase inhibitor combinations, aminoglycosides, fluoroquinolones, tetracycline, glycylcycline, and trimethoprim-sulfamethoxazole. Only one antibiotic resistance gene was recognized in the plasmid. By contrast, many antibiotic resistance genes were identified in the chromosome. The findings of this study suggest that strain ED882-96 is a highly virulent and multidrug-resistant pathogen. Knowledge regarding genomic characteristics and antimicrobial susceptibility patterns provides valuable insights into this uncommon species.

Differences in Clinical Manifestations, Antimicrobial Susceptibility Patterns, and Mutations of Fluoroquinolone Target Genes between Chryseobacterium gleum and Chryseobacterium indologenes.

Chryseobacterium infections are uncommon, and previous studies have revealed that Chryseobacterium gleum is frequently misidentified as Chryseobacterium indologenes We aimed to explore the differences in clinical manifestations and antimicrobial susceptibility patterns between C. gleum and C. indologenes The database of a clinical microbiology laboratory was searched to identify patients with Chryseobacterium infections between 2005 and 2017. Species were reidentified using 16S rRNA gene sequencing, and patients with C. gleum and C. indologenes infections were included in the study. A total of 42 C. gleum and 84 C. indologenes isolates were collected from consecutive patients. A significant increase in C. indologenes incidence was observed. C. gleum was significantly more associated with bacteremia than C. indologenes Patients with C. gleum infections had more comorbidities of malignancy and liver cirrhosis than those with C. indologenes infections. The overall case fatality rate was 19.8%. Independent risk factors for mortality were female sex and C. indologenes infection. These isolates were most susceptible to minocycline (73%), followed by trimethoprim-sulfamethoxazole (47.6%), tigecycline (34.1%), and levofloxacin (32.5%). C. gleum exhibited a significantly higher rate of susceptibility than C. indologenes to piperacillin, piperacillin-tazobactam, ceftazidime, tigecycline, and levofloxacin. Alterations in DNA gyrase subunit A were identified to be associated with fluoroquinolone resistance in C. indologenes No nonsynonymous substitutions were observed in the quinolone resistance-determining regions (QRDRs) of C. gleum Differences in epidemiology, clinical manifestations, and antimicrobial susceptibility patterns exist between C. gleum and C. indologenes Additional investigations are needed to explore the significance of these differences.

Identification of Antibacterial Phenolics in Selected Plant Species from Mexican Cloud Forest by Mass Spectrometry Dereplication.

Fifteen plant species from a protected cloud forest (CF) in Veracruz, Mexico, were screened for their in vitro capacity to inhibit the growth of the phytopathogenic bacteria Chryseobacterium sp., Pseudomonas cichorii, Pectobacterium carotovorum and Pantoea stewartii, causal agents of damage to crops like 'chayote', lettuce, potato and corn. As a result, the bioactivity of Turpinia insignis and Leandra cornoides is reported for the first time against Chryseobacterium sp. and P. cichorii. In addition, 24 and 18 compounds not described for these species were dereplicated by an UPLC/MS-MS method, respectively. The identified compounds included simple phenols, hydroxycinnamic acids, flavonoids and coumarins. The antibacterial assay of 12 of them demonstrated the bacteriostatic effect of vanillin, trans-cinnamic acid, scopoletin and umbelliferone against Chryseobacterium sp. These findings confirm for the first time the value of the CF plants from Veracruz as sources of bioactive natural products with antimicrobial properties against phytopathogenic bacteria.

Elizabethkingia miricola infection in Chinese spiny frog (Quasipaa spinosa).

Elizabethkingia miricola is a Gram-negative rod which has been incriminated in severe infections in humans. Recently, a serious infectious disease was identified in Chinese spiny frogs (Quasipaa spinosa), in the Sichuan Province of China; the disease was characterized by corneal opacity, the presence of ascites and neurological symptoms. A Gram-negative bacillus was isolated from the liver, spleen and kidney of the diseased frogs. Experimental infection test revealed that the bacillus could infect the frogs Q. spinosa and the LD50 value was 1.19 × 106  cfu per frog. The isolated Gram-negative bacillus was identified as E. miricola according to phenotypic characteristics, 16S rRNA and gyrB gene sequence analysis. The isolated strain was only susceptible to florfenicol among all investigated chemotherapeutic agents. Histological examination revealed that E. miricola infection caused pathological lesions to multiple organs and tissues, especially in the liver, brain, kidney. These results confirmed that E. miricola is an emerging pathogen of Chinese spiny frogs.

Molecular Characterization of Chryseobacterium indologenes with Multidrug Resistance in the Brazilian Amazon Region.

Chryseobacterium indologenes is an emerging nosocomial pathogen that produces IND-type chromosomal metallo-beta-lactamase. The phenotype and molecular aspects of two multidrug resistant C. indologenes strains and the analysis of the tertiary structure of the IND enzyme were studied. Identification of species and susceptibility tests were performed using the Vitek-2 compact. Chromosomal and plasmid DNA were extracted using PureLink™ Genomic DNA Mini Kit and PureLink Quick Plasmid Miniprep Kit, and the sequencing was performed using ABI 3130 genetic analyzer. Two strains were isolated and are registered as P-23 and P-113. Of the two, P-113 was sensitive to ciprofloxacin and cefepime only, whereas the P-23 showed reduced sensitivity to ceftazidime, ciprofloxacin, and tigecycline. The genetic analysis of both isolates identified the presence of the blaIND-like gene, with similarity to IND-3 and IND-8 alleles. The IND-3 identified in the P-133 sample presented a single mutation at position T355G, which corresponds to a nonsynonymous substitution of the amino acid at position 119 (Ser→Ala). The phylogenetic analysis of INDs showed lineages that are circulating in Asian and European countries. These results emphasize the need for effective preventive actions to avoid the dissemination of this type of pathogen in the hospital environment.

Treatment considerations for potential uropathogens detected by precision microbiological testing.

PURPOSE: The clinical and microbiological data for urinary tract infections (UTIs) for 6 organisms detected by matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) at community health systems were examined. SUMMARY: The use of precision microbiological diagnostic testing such as MALDI-TOF and real-time quantitative polymerase chain reaction has increased the ability to detect a wider spectrum of organisms. This has raised questions of the clinical relevance of infrequently encountered organisms, especially when cultured from urine. This article reviews clinical and microbiological data for UTIs for 6 organisms detected by MALDI-TOF at community health systems (Actinotignum schaalii, Chryseobacterium indologenes, Aerococcus urinae, Aerococcus sanguinicola, Corynebacterium riegelii, and Corynebacterium urealyticum). Since little information currently exists, most of the data associating the aforementioned organisms with UTIs were derived from case reports. Although these organisms are more readily identified using precision microbiological diagnostic testing methods, infection should not be assumed based on culture results alone since asymptomatic bacteriuria has been reported. Similar to more common urinary pathogens, clinical correlation is essential. To facilitate treatment, we provide a table of empirical options likely to achieve clinical success based on in vivo and in vitro data. If available, pathogen-specific susceptibility data should be used to direct therapy. CONCLUSION: Clinical and microbiological data and potential treatment options were presented for 6 traditionally underrecognized organisms that are increasingly being found from urinary specimens. The treatment recommendations should be interpreted cautiously as they were devised through the use of very limited data.

A rare case of complicated pericardial effusion with Elizabethkingia meningoseptica from Iran.

Infections due to Elizabethkingia meningoseptica, a Gram-negative oxidative bacterium are frequently founded in neonatal and immunocompromised individuals. The notable characteristic of this organism is its multi-drug resistance to common antibiotics used for infections caused by Gram-negative bacteria. We report a rare case of complicated pericardial effusion due to E. meningoseptica in a 2-year-old boy, who was admitted with chief complaints of fever and tachypnea (mentioned by his parents) and suffered from a rare lung malignancy (lymphangioleiomyomatosis). He was successfully treated with vancomycin.  E. meningoseptica infection is a rare situation in immunocompetent hosts, and we concluded that this infection was probably originated from device medicine or even hands of healthcare workers.

Draft genome sequence of Chryseobacterium limigenitum SUR2T (LMG 28734T) isolated from dehydrated sludge

ABSTRACT The type strain SUR2 of the novel species Chryseobacterium limigenitum was isolated from a dehydrated sludge of the municipal sewage treatment plant in Dogoše near Maribor in Slovenia. The draft genome, with 60 contigs, 4,697,725 bp, 34.4% of G+C content, was obtained using the Illumina HiSeq 2500-1 platform. Joint Genome Institute Microbial Genome Annotation Pipeline (MGAP v.4) has identified 4322 protein-coding sequences including resistance genes against arsenic and other heavy metals. In addition, a subclass B3 metallo-β-lactamase, which confers resistance to penicillins, cephalosporins and carbapenems, was also present in the genome. The genome sequence provides important information regarding bioremediation potential and pathogenic properties of this newly identified species.

Polyphasic characterization of bacteria obtained from upland rice cultivated in Cerrado soil

ABSTRACT This work aimed to characterize 20 isolates obtained from upland rice plants, based on phenotypic (morphology, enzymatic activity, inorganic phosphate solubilization, carbon source use, antagonism), genotypic assays (16S rRNA sequencing) and plant growth promotion. Results showed a great morphological, metabolic and genetic variability among bacterial isolates. All isolates showed positive activity for catalase and protease enzymes and, 90% of the isolates showed positive activity for amylase, catalase and, nitrogenase. All isolates were able to metabolize sucrose and malic acid in contrast with mannitol, which was metabolized only by one isolate. For the other carbon sources, we observed a great variability in its use by the isolates. Most isolates showed antibiosis against Rhizoctonia solani (75%) and Sclerotinia sclerotiorum (55%) and, 50% of them showed antibiosis against both pathogens. Six isolates showed simultaneous ability of antibiosis, inorganic phosphate solubilization and protease activity. Based on phylogenetic analysis of the 16S rRNA gene all the isolates belong to Bacillus genus. Under greenhouse conditions, two isolates (S4 and S22) improved to about 24%, 25%, 30% and 31% the Total N, leaf area, shoot dry weight and root dry weight, respectively, of rice plants, indicating that they should be tested for this ability under field conditions.

Polyphasic characterization of bacteria obtained from upland rice cultivated in Cerrado soil.

This work aimed to characterize 20 isolates obtained from upland rice plants, based on phenotypic (morphology, enzymatic activity, inorganic phosphate solubilization, carbon source use, antagonism), genotypic assays (16S rRNA sequencing) and plant growth promotion. Results showed a great morphological, metabolic and genetic variability among bacterial isolates. All isolates showed positive activity for catalase and protease enzymes and, 90% of the isolates showed positive activity for amylase, catalase and, nitrogenase. All isolates were able to metabolize sucrose and malic acid in contrast with mannitol, which was metabolized only by one isolate. For the other carbon sources, we observed a great variability in its use by the isolates. Most isolates showed antibiosis against Rhizoctonia solani (75%) and Sclerotinia sclerotiorum (55%) and, 50% of them showed antibiosis against both pathogens. Six isolates showed simultaneous ability of antibiosis, inorganic phosphate solubilization and protease activity. Based on phylogenetic analysis of the 16S rRNA gene all the isolates belong to Bacillus genus. Under greenhouse conditions, two isolates (S4 and S22) improved to about 24%, 25%, 30% and 31% the Total N, leaf area, shoot dry weight and root dry weight, respectively, of rice plants, indicating that they should be tested for this ability under field conditions.

Draft genome sequence of Chryseobacterium limigenitum SUR2T (LMG 28734T) isolated from dehydrated sludge.

The type strain SUR2 of the novel species Chryseobacterium limigenitum was isolated from a dehydrated sludge of the municipal sewage treatment plant in Dogose near Maribor in Slovenia. The draft genome, with 60 contigs, 4,697,725bp, 34.4% of G+C content, was obtained using the Illumina HiSeq 2500-1 platform. Joint Genome Institute Microbial Genome Annotation Pipeline (MGAP v.4) has identified 4322 protein-coding sequences including resistance genes against arsenic and other heavy metals. In addition, a subclass B3 metallo-ß-lactamase, which confers resistance to penicillins, cephalosporins and carbapenems, was also present in the genome. The genome sequence provides important information regarding bioremediation potential and pathogenic properties of this newly identified species.