Bipartite Genomes in Enterobacterales: Independent Origins of Chromids, Elevated Openness and Donors of Horizontally Transferred Genes.

Multipartite bacteria have one chromosome and one or more chromid. Chromids are believed to have properties that enhance genomic flexibility, making them a favored integration site for new genes. However, the mechanism by which chromosomes and chromids jointly contribute to this flexibility is not clear. To shed light on this, we analyzed the openness of chromosomes and chromids of the two bacteria, Vibrio and Pseudoalteromonas, both which belong to the Enterobacterales order of Gammaproteobacteria, and compared the genomic openness with that of monopartite genomes in the same order. We applied pangenome analysis, codon usage analysis and the HGTector software to detect horizontally transferred genes. Our findings suggest that the chromids of Vibrio and Pseudoalteromonas originated from two separate plasmid acquisition events. Bipartite genomes were found to be more open compared to monopartite. We found that the shell and cloud pangene categories drive the openness of bipartite genomes in Vibrio and Pseudoalteromonas. Based on this and our two recent studies, we propose a hypothesis that explains how chromids and the chromosome terminus region contribute to the genomic plasticity of bipartite genomes.

Salinibacterium sedimenticola sp. nov., Isolated from Tidal Flat Sediment.

An actinobacterium, designated as SYSU T00001T, was isolated from a tidal flat sediment sample from Guangdong province, China. Cells were Gram-stain-positive, aerobic, motile and short rod-shaped. Colonies on marine agar 2216 were smooth, yellow-pigmented, and circular with low convexity. The isolate was able to grow at the temperature range 4-37 °C (optimum 30 °C), at pH 4.0-10.0 (optimum 7.0) and in the presence of 0-10% (w/v) NaCl. The major menaquinones were MK-11 and MK-10. The cell wall contained alanine, glutamic acid, lysine and ornithine. The major fatty acids were C19:0 cyclo ω8c (35.7%) and anteiso C15:0 (26.0%). The polar lipids consisted of one diphosphatidyl glycerol, one unidentified glycolipid and one unknown lipid. Whole genome sequencing of strain SYSU T00001T revealed 2,837,702 bp with a DNA G + C content of 67.8%. Phylogenetic analyses clearly demonstrated that strain SYSU T00001T belonged to the genus Salinibacterium, and the highest 16S rRNA gene similarity to Salinibacterium hongtaonis 194T (97.8%). The ANI and dDDH values of strain SYSU T00001T relative to Salinibacterium hongtaonis 194T were 74.5% and 19.5%, respectively. According to our data, strain SYSU T00001T represents a novel species of the genus Salinibacterium, for which the name Salinibacterium sedimenticola sp. nov. is proposed, the type strain is SYSU T00001T (= GDMCC 1.3283T = KCTC 49758T).

Marinagarivorans cellulosilyticus sp. nov., a cellulolytic bacterium isolated from the deep-sea off Noma-misaki, Japan.

Cells from strain GE09T, isolated from an artificially immersed nanofibrous cellulose plate in the deep sea, were Gram-stain-negative, motile, aerobic cells that could grow with cellulose as their only nutrient. Strain GE09T was placed among members of Cellvibrionaceae, in the Gammaproteobacteria, with Marinagarivorans algicola Z1T, a marine degrader of agar, as the closest relative (97.4 % similarity). The average nucleotide identity and digital DNA-DNA hybridization values between GE09T and M. algicola Z1T were 72.5 and 21.2 %, respectively. Strain GE09T degraded cellulose, xylan and pectin, but not starch, chitin and agar. The different carbohydrate-active enzymes encoded in the genomes of strain GE09T and M. algicola Z1T highlights their differences in terms of target energy sources and reflects their isolation environments. The major cellular fatty acids of strain GE09T were C18 : 1 ω7c, C16 : 0 and C16 : 1 ω7c. The polar lipid profile showed phosphatidylglycerol and phosphatidylethanolamine. The major respiratory quinone was Q-8. Based on these distinct taxonomic characteristics, strain GE09T represents a new species in the genus Marinagarivorans, for which we propose the name Marinagarivorans cellulosilyticus sp. nov. (type strain GE09T=DSM 113420T=JCM 35003T).

A global perspective on bacterial diversity in the terrestrial deep subsurface.

While recent efforts to catalogue Earth's microbial diversity have focused upon surface and marine habitats, 12-20 % of Earth's biomass is suggested to exist in the terrestrial deep subsurface, compared to ~1.8 % in the deep subseafloor. Metagenomic studies of the terrestrial deep subsurface have yielded a trove of divergent and functionally important microbiomes from a range of localities. However, a wider perspective of microbial diversity and its relationship to environmental conditions within the terrestrial deep subsurface is still required. Our meta-analysis reveals that terrestrial deep subsurface microbiota are dominated by Betaproteobacteria, Gammaproteobacteria and Firmicutes, probably as a function of the diverse metabolic strategies of these taxa. Evidence was also found for a common small consortium of prevalent Betaproteobacteria and Gammaproteobacteria operational taxonomic units across the localities. This implies a core terrestrial deep subsurface community, irrespective of aquifer lithology, depth and other variables, that may play an important role in colonizing and sustaining microbial habitats in the deep terrestrial subsurface. An in silico contamination-aware approach to analysing this dataset underscores the importance of downstream methods for assuring that robust conclusions can be reached from deep subsurface-derived sequencing data. Understanding the global panorama of microbial diversity and ecological dynamics in the deep terrestrial subsurface provides a first step towards understanding the role of microbes in global subsurface element and nutrient cycling.

Transcriptionally active nasopharyngeal commensals and opportunistic microbial dynamics define mild symptoms in the COVID 19 vaccination breakthroughs.

The development of COVID 19 vaccines as an effort to mitigate the outbreak, has saved millions of lives globally. However, vaccination breakthroughs have continuously challenged the vaccines' effectiveness and provided incentives to explore facets holding potential to alter vaccination-induced immunity and protection from subsequent infection, especially VOCs (Variants Of Concern). We explored the functional dynamics of nasopharyngeal transcriptionally active microbes (TAMs) between vaccination breakthroughs and unvaccinated SARS-CoV-2 infected individuals. Microbial taxonomic communities were differentially altered with skewed enrichment of bacterial class/genera of Firmicutes and Gammaproteobacteria with grossly reduced phylum Bacteroidetes in vaccination breakthrough individuals. The Bacillus genus was abundant in Firmicutes in vaccination breakthrough whereas Prevotella among Bacteroides dominated the unvaccinated. Also, Pseudomonas and Salmonella of Gammaproteobacteria were overrepresented in vaccination breakthrough, whilst unvaccinated showed presence of several genera, Achromobacter, Bordetella, Burkholderia, Neisseria, Hemophilus, Salmonella and Pseudomonas, belonging to Proteobacteria. At species level, the microbiota of vaccination breakthrough exhibited relatively higher abundance of unique commensals, in comparison to potential opportunistic microbes enrichment in unvaccinated patients' microbiota. Functional metabolic pathways like amino acid biosynthesis, sulphate assimilation, fatty acid and beta oxidation, associated with generation of SCFAs (short chain fatty acids), were enriched in vaccination breakthroughs. Majorly, metabolic pathways of LCFAs biosynthesis (long chain fatty acids; oleate, dodecenoate, palmitoleate, gondoate) were found associated with the unvaccinated. Our research highlights that vaccination decreases the microbial diversity in terms of depleting opportunistic pathogens and increasing the preponderance of commensals with respect to unvaccinated patients. Metabolic pathway analysis substantiates the shift in diversity to functionally modulate immune response generation, which may be related to mild clinical manifestations and faster recovery times during vaccination breakthroughs.

Identification and characterization of a novel high-activity amylosucrase from Salinispirillum sp. LH10-3-1.

In this study, a novel high-activity amylosucrase from Salinispirillum sp. LH10-3-1 (SaAS) was identified and characterized. The recombinant enzyme was determined as a monomer with a molecular mass of 75 kDa. SaAS protein exhibited the maximum total and polymerization activities at pH 9.0 and maximum hydrolysis activity at pH 8.0. The optimum temperature for total, polymerization, and hydrolysis activities were 40, 40, and 45 °C, respectively. Under the optimal pH and temperature, SaAS had a specific activity of 108.2 U/mg. SaAS also showed excellent salt tolerance and could retain 77.4% of its original total activity at 4.0 M NaCl. The addition of Mg2+, Ba2+, and Ca2+ enhanced the total activity of SaAS. When the conversion of 0.1 M and 1.0 M sucrose was catalyzed at pH 9.0 and 40 °C for 24 h, the ratios of hydrolysis, polymerization, and isomerization reactions were 11.9:77.4:10.7 and 15.3:53.5:31.2, respectively. The α-arbutin yield of 60.3% was achieved from 20 mM sucrose and 5 mM hydroquinone catalyzed by SaAS. KEY POINTS: • A novel amylosucrase from Salinispirillum sp. LH10-3-1 (SaAS) was characterized. • SaAS has the highest specific enzyme activity among all known amylosucrase. • SaAS has hydrolysis, polymerization, isomerization, and glucosyltransferase activities.

Chitinophaga horti sp. nov., Isolated from Garden Soil.

A strictly aerobic Gram-negative bacterium, designated R8T, isolated from garden soil in South Korea was subjected to a taxonomic study. The cells were non-spore-forming, oxidase-positive and catalase-negative, and non-motile rods (without flagella). Growth was observed between 10 °C and 40 °C (optimum, 30 °C) and between pH 6.0 and 9.0 (optimum, pH 7.0) and in the presence of 0%-1.5% (w/v) NaCl (optimum, 0%). The G + C content of the genomic DNA was 49.9% and the major isoprenoid quinone was found to be menaquinone-7. The major fatty acids of strain R8T were iso-C15:0, C16:1 ω5c, and summed feature 3 (comprising iso-C15:0 2-OH and/or C16:1 ω7c/ω6c). Phosphatidylethanolamine was identified as a major polar lipid. Comparative 16S rRNA gene sequence analysis showed that strain R8T had the highest 16S rRNA gene sequence similarity of 98.3% with Chitinophaga sedimenti TFL-3 T. Phylogenetic analyses using 16S rRNA gene sequences and concatenated 92 marker protein sequences revealed that strain R8T formed a robust phylogenetic lineage with C. sedimenti within the genus Chitinophaga. Average nucleotide identity and digital DNA-DNA hybridization values of strain R8T to Chitinophaga species were less than 77.9% and 21.1%, respectively. The phenotypic, phylogenetic, and chemotaxonomic properties support that strain R8T represents a novel species of the genus Chitinophaga, for which the name Chitinophaga horti sp. nov. is proposed. The type strain is R8T (= KACC 19895 T = JCM 33215 T).

Proposal of Holzapfeliella gen. nov. and Litorivicinus gen. nov. as replacement names for the illegitimate prokaryotic generic names Holzapfelia Zheng et al. 2020 and Litoricola Kim et al. 2007, respectively.

The prokaryotic generic names Holzapfelia Zheng et al. 2020 and Litoricola Kim et al. 2007 are illegitimate because they are later homonyms of the names Holzapfelia Cossmann 1901, an extinct genus of molluscs (Neogastropoda) and Litoricola Woodward 1873, an extinct genus of crustaceans (Arthropoda), respectively (Principle 2 and Rule 51b(4) of the International Code of Nomenclature of Prokaryotes). We therefore propose the replacement generic name Holzapfeliella with type species Holzapfeliella floricola comb. nov. and the replacement generic name Litorivicinus with replacement name Litorivicinus lipolyticus for the type species, the replacement name Litorivicinus marinus, and the family name Litorivicinaceae to replace the illegitimate name Litoricolaceae Kim et al. 2007.

Community assemblages and species coexistence of prokaryotes controlled by local environmental heterogeneity in a cold seep water column.

The distribution and heterogeneity characteristics of microbial communities in cold seep water columns are significant factors governing the efficiency of methane filtering and carbon turnover. However, this process is poorly understood. The diversity of vertically stratified microbial communities and the factors controlling the community assemblage process in the water column above the Haima cold seep were investigated in this study. The prokaryotic community diversities varied distinctly with vertical changes in hydrochemistry. Cyanobacteria dominated the light-transmitting layers and Proteobacteria dominated the deeper layers. With respect to microbial community assemblages and co-occurrence networks, stochastic processes were particularly important in shaping prokaryotic communities. In the shallow (≥85 m) and mesopelagic water columns (600-800 m), microbial community characteristics were affected by deterministic processes, reduced network connectivity, and modularity. Microbial community diversities and assemblage processes along a vertical profile were influenced by the vertical variations in pH, temperature, DIC, and nutrients. Stochastic processes may have facilitated the formation of complex co-occurrence networks. Briefly, the distribution of local environmental heterogeneity along the vertical dimension could drive unique microbial community assemblage and species coexistence patterns. This study provides new perspectives on how microorganisms adapt to the environment and build communities, and how species coexist in shared habitats.

Bacterial response to glucose addition: growth and community structure in seawater microcosms from North Pacific Ocean.

Onboard microcosm experiments were conducted to assess how bacterial growth pattern and community structure changed by the addition of labile organic compound during the KH-14-2 cruise of R/V Hakuho Maru (Atmosphere and Ocean Research Institute, the University of Tokyo and JAMSTEC) in May-June 2014. Seawater samples were collected from the three diversified oceanic environments, Kuroshio Current, North Pacific Sub-polar Gyre (SPG), and North Pacific Sub-tropical Gyre (STG) in the western North Pacific Ocean, filtered, supplemented with glucose, and incubated at 23 ± 1 °C, ~ 4 °C, and 23 ± 1 °C, respectively. Untreated control microcosms were also maintained for all the sample types. Significant increases in cell counts and cell sizes were observed in Kuroshio Current and STG waters, whereas in SPG neither the counts nor the sizes changed, even after 120 h of incubation. At early stages of incubation, the classes Bacteroidia, Alphaproteobacteria, and Gammaproteobacteria were dominant in the Kuroshio Current and SPG samples, while the phyla Cyanobacteria and Proteobacteria in the STG samples. Over incubation periods between 60 and 96 h, some members of the class Gammaproteobacteria gradually dominated within which the genera Vibrio and Alteromonas became dominant in the Kuroshio Current and STG, respectively. No growth was detected for the microcosms with seawater from SPG, regardless of glucose amendment. It is concluded that depending on the environmental condition, certain different bacterial groups proliferated quickly and modified the community structures. Temperature significantly influenced the growth and succession, and ultimately the community structure of bacteria.

Intestinal colonization with multidrug-resistant Enterobacterales: screening, epidemiology, clinical impact, and strategies to decolonize carriers.

The clinical impact of infections due to extended-spectrum ß-lactamase (ESBL)- and/or carbapenemase-producing Enterobacterales (Ent) has reached dramatic levels worldwide. Infections due to these multidrug-resistant (MDR) pathogens-especially Escherichia coli and Klebsiella pneumoniae-may originate from a prior asymptomatic intestinal colonization that could also favor transmission to other subjects. It is therefore desirable that gut carriers are rapidly identified to try preventing both the occurrence of serious endogenous infections and potential transmission. Together with the infection prevention and control countermeasures, any strategy capable of effectively eradicating the MDR-Ent from the intestinal tract would be desirable. In this narrative review, we present a summary of the different aspects linked to the intestinal colonization due to MDR-Ent. In particular, culture- and molecular-based screening techniques to identify carriers, data on prevalence and risk factors in different populations, clinical impact, length of colonization, and contribution to transmission in various settings will be overviewed. We will also discuss the standard strategies (selective digestive decontamination, fecal microbiota transplant) and those still in development (bacteriophages, probiotics, microcins, and CRISPR-Cas-based) that might be used to decolonize MDR-Ent carriers.

Comparison of the relative efficacy of ß-lactam/ß-lactamase inhibitors and carbapenems in the treatment of complicated urinary tract infections caused by ceftriaxone-non-susceptible Enterobacterales: a multicentre retrospective observational cohort study.

BACKGROUND: Treating complicated urinary tract infections (cUTIs) caused by ESBL-producing Enterobacterales represents a significant clinical challenge. The present study was thus developed to explore the relative efficacy of ß-lactam/ß-lactamase inhibitors (BLBLIs) and carbapenems for the treatment of hospitalized patients suffering from cUTIs caused by BLBLI-susceptible ceftriaxone-non-susceptible Enterobacterales. METHODS: Data from 557 patients from four Chinese teaching hospitals diagnosed with cUTIs caused by ceftriaxone-non-susceptible Enterobacterales from January 2017 to May 2022 were retrospectively assessed. RESULT: The 30 day rate of treatment failure, defined by unresolved symptoms or mortality, was 10.4% (58/557). Independent predictors of 30 day treatment failure included immunocompromised status, bacteraemia, septic shock, lack of infection source control and appropriate empirical treatment. When data were controlled for potential confounding variables, BLBLI treatment exhibited a comparable risk of 14 day (OR 1.61, 95% CI 0.86-3.00, P = 0.133) and 30 day treatment failure (OR 1.45, 95% CI 0.66-3.15, P = 0.354) relative to carbapenem treatment for the overall cohort of patients. In contrast, BLBLI treatment in immunocompromised patients was associated with an elevated risk of both 14 day (OR 3.18, 95% CI 1.43-7.10, P = 0.005) and 30 day treatment failure (OR 3.06, 95% CI 1.07-8.80, P = 0.038) relative to carbapenem treatment. CONCLUSIONS: These results suggested that carbapenem treatment may be superior to BLBLI treatment for immunocompromised patients suffering from cUTIs caused by ceftriaxone-non-susceptible Enterobacterales species. However, these results will need to be validated in appropriately constructed randomized controlled trials to ensure appropriate patient treatment.

Genomic analysis of Marinimicrobium sp. C6131 reveals its genetic potential involved in chitin metabolism.

Marinimicrobium sp. C6131, which had the ability to degrade chitin, was isolated from deep-sea sediment of the southwest Indian Ocean. Here, the genome of strain C6131 was sequenced and the chitin metabolic pathways were constructed. The genome contained a circular chromosome of 4,207,651 bp with a G + C content of 58.50%. A total of 3471 protein-coding sequences were predicted. Gene annotation and metabolic pathway reconstruction showed that strain C6131 possessed genes and two metabolic pathways involved in chitin catabolism: the hydrolytic chitin utilization pathway initiated by chitinases and the oxidative chitin utilization pathway initiated by lytic polysaccharide monooxygenases. Chitin is the most abundant polysaccharide in the ocean. Degradation and recycling of chitin driven by marine bacteria are crucial for biogeochemical cycles of carbon and nitrogen in the ocean. The genomic information of strain C6131 revealed its genetic potential involved in chitin metabolism. The strain C6131 could grow with colloidal chitin as the sole carbon source, indicating that these genes would have functions in chitin degradation and utilization. The genomic sequence of Marinimicrobium sp. C6131 could provide fundamental information for future studies on chitin degradation, and help to improve our understanding of the chitin degradation process in deep-sea environments.

Diversity and Quantitative Detection of Clade I Type nosZ Denitrifiers in the Arabian Sea Oxygen Minimum Zone.

A significant amount of nitrous oxide (N2O) is effluxed into the atmosphere as a result of marine denitrification in the Arabian Sea (AS) oxygen minimum zone (OMZ). An assessment of temporal variations in the diversity and abundance of nosZ denitrifiers was performed to establish the relative importance of these bacteria in denitrification. Sampling was conducted at the Arabian Sea Time Series (ASTS) location and a quantitative PCR (qPCR) ana-lysis was performed. We detected a high abundance of the nosZ gene at core OMZ depths (250| |m and 500 m), indicating the occurrence of denitrification in the AS-OMZ. The maximum abundance of the nosZ gene was observed during the Spring Intermonsoon (SIM) at 250| |m (1.32×106 copies L-1) and 500| |m (1.50×106 copies L-1). Sequencing ana-lysis showed that nosZ denitrifiers belonged to the classes Alpha-, Beta-, and Gammaproteobacteria. Taxonomic ana-lysis revealed that most OTUs were affiliated with Pseudomonas, Rhodopseudomonas, and Bradyrhizobium. Diversity indices and richness estimators confirmed a higher diversity of nosZ denitrifiers at 250| |m than at 500| |m during all three seasons. The present results also indicated that dissolved oxygen (DO) and total organic carbon (TOC) are critical factors influencing the diversity and abundance of the nosZ-denitrifying bacterial community.

Phylogenetic group-based assembly and co-occurrence pattern of the microbial community in full-scale wastewater treatment plants during the Chinese spring festival.

The quality and quantity of domestic sewage discharge vary significantly during the Chinese Spring Festival due to the huge population shift. The dynamics of microbial community traits during the Spring Festival, particularly the phylogenetic group-based assembly and co-occurrence patterns, are however little understood. Here, influent and activated sludge samples from 2 full-scale wastewater treatment plants were collected bi-daily throughout a 20-day Spring Festival period and subjected to high-throughput Illumina-MiSeq sequencing. The findings revealed that the microbial communities in the activated sludge displayed a comparatively stable pattern, and that the influent communities experienced significant temporal fluctuations in terms of diversity and composition. The characterization by "Infer Community Assembly Mechanisms by Phylogenetic-bin based null model" demonstrated that for Competibacter glycogen-accumulating organisms, the assembly mechanism shifted from deterministic process (HoS = 69.5%) before the Spring Festival to stochastic process (DR = 65.9%) after the Spring Festival. The network analysis revealed that the network structure of sludge communities was more stable before the Spring Festival than that after the Spring Festival. Additionally, sludge communities had no keystone species in common with the influent before the Spring Festival, while the sludge and influent communities shared two keystone taxa after the Spring Festival (Sebaldella and Candidatus Competibacter). This study would deepen our understanding of the microbial ecology in biological wastewater treatment systems, which also aids in managing wastewater treatment plants.

In Vitro Activity of Ceftibuten-Avibactam against ß-Lactamase-Positive Enterobacterales from the ATLAS Global Surveillance Program.

Ceftibuten is an established, oral, third-generation cephalosporin in early clinical development in combination with an oral prodrug of avibactam for the treatment of complicated urinary tract infections, including acute pyelonephritis. We evaluated the in vitro activity of ceftibuten-avibactam against 1,165 Enterobacterales isolates selected from the 2016-2020 ATLAS global surveillance program based upon their ß-lactamase genotype, ß-lactam-susceptible phenotype, species identification, and specimen source (95.8% urine). MICs were determined by CLSI broth microdilution. Avibactam was tested at a fixed concentration of 4 µg/mL. Molecular methods were used to identify ß-lactamase genes. Ceftibuten-avibactam inhibited 90% (MIC90) of ESBL-producing (n = 645), KPC-producing (n = 60), chromosomal AmpC-positive (n = 100), OXA-48-like-producing (n = 50), and acquired AmpC-producing (n = 110) isolates at concentrations of 0.12, 0.5, 1, 2, and 4 µg/mL, respectively. At concentrations of ≤1 and ≤8 µg/mL, ceftibuten-avibactam inhibited 98.4 and 99.2% of ESBL-positive isolates; 96.7 and 100% of KPC-positive isolates; 91.0 and 99.0% of chromosomal AmpC-positive isolates; 86.0 and 96.0% of OXA-48-like-positive isolates; and 85.5 and 91.8% of acquired AmpC-positive isolates. Against ESBL-producing, KPC-producing, chromosomal AmpC-positive, OXA-48-like-producing, and acquired AmpC-producing isolates, ceftibuten-avibactam was 256-, 128-, >64-, >32-, and > 16-fold more potent than ceftibuten alone. The potency of ceftibuten-avibactam was 4-fold greater than ceftazidime-avibactam against ESBL-producing (ceftibuten-avibactam MIC90, 0.12 µg/mL; ceftazidime-avibactam MIC90, 0.5 µg/mL) and KPC-producing (0.5 µg/mL; 2 µg/mL) isolates, equivalent to ceftazidime-avibactam (MIC90, 2 µg/mL) against OXA-48-like-producing isolates, 2-fold less active than ceftazidime-avibactam (1 µg/mL; 0.5 µg/mL) against chromosomal AmpC-positive isolates, and 4-fold less active than ceftazidime-avibactam (4 µg/mL; 1 µg/mL) against acquired AmpC-producing isolates. Continued development of ceftibuten-avibactam appears justified.

Meropenem-Vaborbactam Activity against U.S. Multidrug-Resistant Enterobacterales Strains, Including Carbapenem-Resistant Isolates.

Carbapenems are a common first-line therapy for serious Gram-negative infections, but carbapenem-resistant Enterobacterales (CRE) isolates have become an urgent health concern. Klebsiella pneumoniae serine carbapenemases (KPCs) now have been disseminated worldwide and are endemic in many hospitals globally. Isolates producing metallo-ß-lactamases (MBLs) or class D OXA-48 carbapenemases are also increasingly common in Europe, although they are less common in the United States. Meropenem-vaborbactam is a combination of the carbapenem meropenem and vaborbactam, which is a ß-lactamase inhibitor with activity against serine carbapenemases, including KPC-producing isolates. We examined the susceptibility of U.S. multidrug-resistant (MDR) isolates to meropenem-vaborbactam. A total of 1,697 MDR Enterobacterales isolates were collected in 31 U.S. medical centers in 2016 to 2020. Susceptibility testing was performed using the Clinical and Laboratory Standards Institute (CLSI) broth microdilution method. Whole-genome sequencing was performed for all CRE strains (MIC values of >2 mg/L for imipenem or meropenem). The rate of susceptibility of all MDR Enterobacterales strains to meropenem-vaborbactam was 99.1%, and 86.2% of the isolates were susceptible to meropenem. There were 222 CRE isolates (13.1%). KPC was the most common carbapenemase (81.1%). Thirteen CRE isolates produced NDM (n = 7), VIM (n = 3), and/or OXA-48-like (n = 4) carbapenemases; 29 CRE isolates (13.1%) had no detected carbapenemase. The rate of susceptibility of all CRE strains to meropenem-vaborbactam was 93.2%, and the rate of susceptibility of the KPC-producing isolates to meropenem-vaborbactam was 98.9%. The primary carbapenemase in the United States continues to be KPC, while MBL and OXA-48-like carbapenemases remain uncommon. Overall, the rate of susceptibility of these U.S. MDR organisms to meropenem-vaborbactam was 99.1%, indicating that meropenem-vaborbactam is a valuable treatment option for Gram-negative infections caused by U.S. MDR organisms. IMPORTANCE Carbapenems are a common first-line therapy for serious Gram-negative infections, but CRE isolates have become an urgent health concern. Meropenem-vaborbactam is a combination of the carbapenem meropenem and vaborbactam, which is a ß-lactamase inhibitor with activity against serine carbapenemases, including KPC-producing isolates. We examined the susceptibility of U.S. MDR Gram-negative isolates to meropenem-vaborbactam. A total of 1,697 U.S. MDR Enterobacterales isolates collected in 2016 to 2020 were tested. Susceptibility testing was performed using the CLSI broth microdilution method. Whole-genome sequencing was performed for all CRE strains (MIC values of >2 mg/L for imipenem or meropenem). The rate of susceptibility of all MDR Enterobacterales strains to meropenem-vaborbactam was 99.1%, and 86.2% of the isolates were susceptible to meropenem. A total of 13.1% of the isolates were CRE strains, and KPC was the most common carbapenemase. Overall, the rate of susceptibility of these U.S. MDR organisms to meropenem-vaborbactam indicates that meropenem-vaborbactam is a valuable treatment option for Gram-negative infections caused by U.S. MDR Gram-negative pathogens.

Longitudinal Study of Dynamic Epidemiology of Extended-Spectrum Beta-Lactamase-Producing Escherichia coli in Pigs and Humans Living and/or Working on Pig Farms.

Extended-spectrum beta-lactamase (ESBL)-producing Enterobacterales have been increasingly isolated from pigs, highlighting their potential for transmission to humans living and/or working within pig farms. As longitudinal data on the prevalence and the molecular characteristics of such isolates from the high-risk farming population remain scarce, we performed a long-term study on 39 Dutch pig farms. Fecal samples from pigs, farmers, family members, and employees were collected during four sampling occasions with a 6-month period. The presence of ESBL-producing Enterobacterales and their molecular characteristics (ESBL gene, plasmid, and sequence types) were determined by standard methods. Data on personal and farm characteristics were collected using questionnaires. ESBL-producing Escherichia coli was present in pigs at least once for 18 of 39 farms and in 17 of 146 farmers, family members, and/or employees. Among these 417 E. coli isolates, blaCTX-M-1 was the most frequently observed ESBL gene in pigs (n = 261) and humans (n = 25). Despite the great variety in plasmid (sub)types and E. coli sequence types (STs), we observed genetic similarity between human- and pig-derived isolates in (i) ESBL gene, plasmid (sub)type, and ST, suggesting potential clonal transmission in seven farms, and (ii) only ESBL gene and plasmid (sub)type, highlighting the possibility of horizontal transfer in four farms. Five pig farmers carried ESBL producers repeatedly, of whom two carried an identical combination of gene, plasmid (sub)type, and ST over time. Human ESBL carriage was associated with both presence of ESBL producers in pigs and average number of hours working on the pig farm per week, while prolonged human carriage was observed only incidentally. IMPORTANCE Extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli represents a public health hazard due to reduced therapeutic options for the treatment of infections. Although direct contact with pigs is considered a risk factor for human ESBL-producing E. coli carriage through occupational exposure, nationwide data regarding the occurrence of such isolates among pigs and humans living and/or working on farms remain scarce. Therefore, we determined (i) the longitudinal dynamics in prevalence and molecular characteristics of ESBL-producing E. coli in Dutch pig farmers and their pigs over time and (ii) the potential transmission events between these reservoirs based on genetic relatedness and epidemiological associations in longitudinal data. Our data suggesting the possibility of clonal and horizontal dissemination of ESBL-producing Escherichia coli between pigs and pig farmers can be used to inform targeted intervention strategies to decrease the within-farm human exposure to ESBL-producing E. coli.

QMAC-dRAST for the direct testing of antibiotic susceptibility for Enterobacterales in positive blood-culture broth: a comparison of the performances with the MicroScan system and direct disc diffusion testing methods.

OBJECTIVES: To evaluate the performances of the QMAC-dRAST GN (Gram-negative) kit for rapid antimicrobial sensitivity testing (AST) and two other methods, directly on positive blood-culture broth (PBCB), by comparison with a reference method: the MicroScan method based on broth microdilution on colonies isolated on PBCB subculture. METHODS: In total, 156 samples were collected prospectively from blood cultures positive for a Gram-negative rod. Each sample was tested with four AST techniques: (i) the QMAC dRAST GN kit, (ii) the disc diffusion (DD) method, (iii) the MicroScan method applied directly to PBCB; and (iv) MicroScan with isolates from PBCB subculture, as a reference. RESULTS: For 124 PBCB containing Enterobacterales, overall essential agreement (EA) and categorical agreement (CA) between the QMAC-dRAST on PBCB and the reference reached 95.7% and 93.5%, respectively. There were 3.0% very major errors (VME), 4.0% major errors (ME) and 2.8% minor errors (mE). A comparison of MicroScan on PBCB and the reference yielded 98.8% EA, 98.5% CA, and rates of 0.6% VME, 0.9% ME and 0.7% mE. The DD method on PBCB gave a CA of 95.8% and rates of 1.7% for VME, 2.0% for ME and 1.9% for mE. Results were obtained more rapidly for QMAC-dRAST (median of 6 h 37 min versus 18 h for the MicroScan and DD methods on PBCB). CONCLUSIONS: The QMAC-dRAST system provided rapid results well correlated with the reference method on PBCB containing Enterobacterales. Given the shorter time-to-results, the QMAC-dRAST system constitutes a fast and reliable alternative to conventional AST methods.

Diversity dynamics of aerobic anoxygenic phototrophic bacteria in a freshwater lake.

Aerobic anoxygenic photoheterotrophic (AAP) bacteria represent a functional group of prokaryotic organisms that harvests light energy using bacteriochlorophyll-containing photosynthetic reaction centers. They represent an active and rapidly growing component of freshwater bacterioplankton, with the highest numbers observed usually in summer. Species diversity of freshwater AAP bacteria has been studied before in lakes, but its seasonal dynamics remain unknown. In this report, we analysed temporal changes in the composition of the phototrophic community in an oligo-mesotrophic freshwater lake using amplicon sequencing of the pufM marker gene. The AAP community was dominated by phototrophic Gammaproteobacteria and Alphaproteobacteria, with smaller contribution of phototrophic Chloroflexota and Gemmatimonadota. Phototrophic Eremiobacteriota or members of Myxococcota were not detected. Interestingly, some AAP taxa, such as Limnohabitans, Rhodoferax, Rhodobacterales or Rhizobiales, were permanently present over the sampling period, while others, such as Sphingomonadales, Rhodospirillales or Caulobacterales appeared only transiently. The environmental factors that best explain the seasonal changes in AAP community were temperature, concentrations of oxygen and dissolved organic matter.