A Multiwell-Plate Caenorhabditis elegans Assay for Assessing the Therapeutic Potential of Bacteriophages against Clinical Pathogens.

In order to establish phage therapy as a standard clinical treatment for bacterial infections, testing of every phage to ensure the suitability and safety of the biological compound is required. While some issues have been addressed over recent years, standard and easy-to-use animal models to test phages are still rare. Testing of phages in highly suitable mammalian models such as mice is subjected to strict ethical regulations, while insect larvae such as the Galleria mellonella model suffer from batch-to-batch variations and require manual operator skills to inject bacteria, resulting in unreliable experimental outcomes. A much simpler model is the nematode Caenorhabditis elegans, which feeds on bacteria, a fast growing and easy to handle organism that can be used in high-throughput screening. In this study, two clinical bacterial strains of Escherichia coli, one Klebsiella pneumoniae, and one Enterobacter cloacae strain were tested on the model system together with lytic bacteriophages that we isolated previously. We developed a liquid-based assay, in which the efficiency of phage treatment was evaluated using a scoring system based on microscopy and counting of the nematodes, allowing increasing statistical significance compared to other assays such as larvae or mice. Our work demonstrates the potential to use Caenorhabditis elegans to test the virulence of strains of Klebsiella pneumoniae, Enterobacter cloacae, and EHEC/EPEC as well as the efficacy of bacteriophages to treat or prevent infections, allowing a more reliable evaluation for the clinical therapeutic potential of lytic phages. IMPORTANCE Validating the efficacy and safety of phages prior to clinical application is crucial to see phage therapy in practice. Current animal models include mice and insect larvae, which pose ethical or technical challenges. This study examined the use of the nematode model organism C. elegans as a quick, reliable, and simple alternative for testing phages. The data show that all the four tested bacteriophages can eliminate bacterial pathogens and protect the nematode from infections. Survival rates of the nematodes increased from <20% in the infection group to >90% in the phage treatment group. Even the nematodes with poly-microbial infections recovered during phage cocktail treatment. The use of C. elegans as a simple whole-animal infection model is a rapid and robust way to study the efficacy of phages before testing them on more complex model animals such as mice.

Formation, collective motion, and merging of macroscopic bacterial aggregates.

Chemotactic bacteria form emergent spatial patterns of variable cell density within cultures that are initially spatially uniform. These patterns are the result of chemical gradients that are created from the directed movement and metabolic activity of billions of cells. A recent study on pattern formation in wild bacterial isolates has revealed unique collective behaviors of the bacteria Enterobacter cloacae. As in other bacterial species, Enterobacter cloacae form macroscopic aggregates. Once formed, these bacterial clusters can migrate several millimeters, sometimes resulting in the merging of two or more clusters. To better understand these phenomena, we examine the formation and dynamics of thousands of bacterial clusters that form within a 22 cm square culture dish filled with soft agar over two days. At the macroscale, the aggregates display spatial order at short length scales, and the migration of cell clusters is superdiffusive, with a merging acceleration that is correlated with aggregate size. At the microscale, aggregates are composed of immotile cells surrounded by low density regions of motile cells. The collective movement of the aggregates is the result of an asymmetric flux of bacteria at the boundary. An agent-based model is developed to examine how these phenomena are the result of both chemotactic movement and a change in motility at high cell density. These results identify and characterize a new mechanism for collective bacterial motility driven by a transient, density-dependent change in motility.

Population-based surveillance of Enterobacter cloacae complex causing blood stream infections in a centralized Canadian region.

Active population-based surveillance determined clinical factors, susceptibility patterns, incidence rates (IR), and genomics among Enterobacter cloacae complex (n = 154) causing blood stream infections in a centralized Canadian region (2015-2017). The annual population IR was 1.2/100,000 (95% CI 0.9-16) in 2015, 1.4/100,000 (95% CI 1.1-1.9) in 2016, and 1.5/100,000 (95% CI 1.2-2.0) in 2017, affecting mainly elderly males with underlying comorbid conditions in the hospital setting. E. cloacae complex was dominated by polyclonal subspecies (i.e., E. hormaechei subsp. steigerwaltii, subsp. hoffmanni and subsp. xiangfangesis). Antimicrobial resistant determinants were rare. This study provided novel information about Enterobacter genomics in a well-defined human population.

Scavenger receptor B1 mediates phagocytosis and the antimicrobial peptide pathway in the endoparasitic wasp Micropilits mediator.

Scavenger receptors (SRs) are a family of pattern recognition receptors (PRRs) in the immune system. They are required for phagocytosis and act as co-receptors of Toll-like receptors to regulate immune signaling pathways in the fight against pathogens. Little is known about the function of SRs in insects. Here, we reported on a member of the SR family from the parasitic wasp Micropilits mediator (designated MmSR-B1) that is responsive to bacterial infection. The recombinant extracellular CD36 domain of MmSR-B1 produced in Escherichia coli cells is capable of binding to peptidoglycans and bacterial cells, causing agglutination of bacteria. Furthermore, we demonstrated that double-stranded RNA-mediated knockdown of MmSR-B1 impedes hemocyte phagocytosis and downregulates the expression of antimicrobial peptide (AMP) genes defensins and hymenoptaecins. Knockdown of MmSR-B1 led to increased death of the wasps when challenged by bacteria. Our study suggests that MmSR-B1 mediates phagocytosis and the production of AMPs in M. mediator wasps.

Transcriptome analysis of the innate immune system of Hyalomma asiaticum.

Ticks are considered to be the second most important vectors of human infectious diseases. The innate immune system is the key factor that affects its vector competence. Hyalomma asiaticum is the primary vector of Crimean-Congo hemorrhagic fever virus (CCHFV). However, the immune system of H. asiaticum remains virtually unknown. Here, a high throughput full-length mRNA sequencing method was adopted to define the immunotranscriptome of H. asiaticum infected with the fungal pathogen Beauveria bassiana and gram-negative bacterium Enterobacter cloacae. The analysis yielded 22,300 isoforms with an average length of 3233 bps. In total, 68 potential immunity-related genes were identified based on similarity to the homologs known to be involved in immunity. These included most members of the Toll and JAK/STAT signaling pathways, but not the IMD signaling pathway. Moreover, two copies of Dicer-2 and five copies of Argonaute-2 were detected. These genes are postulated to be involved in the RNA interference (RNAi) pathway, which is an important defense against RNA viruses. Overall, this study provides the foundation for understanding the immune response of H. asiaticum to CCHFV.

Mosquito midgut Enterobacter cloacae and Serratia marcescens affect the fitness of adult female Anopheles gambiae s.l.

Some bacteria species found in the mosquito midgut have demonstrated their role in interrupting the development of Plasmodium within the midgut of the Anopheles mosquito and have been identified as potential candidates for novel bacteria-mediated disease control. However, to use these bacteria successfully in biocontrol mechanisms their effect on the fitness of the vector into which they have been introduced has to be evaluated. This study investigated the effect of two such bacteria candidates, Enterobacter cloacae and Serratia marcescens, on Anopheles gambiae s.l. fitness. Pupae and larvae of Anopheles gambiae s.l. mosquitoes were collected by dipping method and reared to adults. The effect of these bacteria on mosquito fitness was assessed by reintroducing isolates of each bacteria separately into antibiotic-treated female adult mosquitoes through sugar meal. Wild type (non-antibiotic-treated) mosquitoes and those antibiotic-treated with no bacteria reintroduction were used as controls. The mosquitoes were monitored on longevity/survival, fecundity, hatch rate, and larval survival. The antibiotic-treated adult mosquitoes had reduced life span with median survival of 14 days while the bacteria-reintroduced groups and the wild type survived to day 22 (p< 0.0001). Treatment with Enterobacter and Serratia did not affect the average egg deposition (p>0.05) but they affected hatch rates positively (p = 0.008). There was, however, some evidence that suggests Enterobacter could have a positive effect on larval development (p < 0.0001). With no observed negative effect on survival/longevity of Anopheles gambiae, introducing E. cloacae and S. marcescens in future bacteria-associated control strategies is unlikely to result in mosquitoes that will be outlived by the wild population. This, however, requires evaluations under field conditions.

Genome-wide Analysis of Four Enterobacter cloacae complex type strains: Insights into Virulence and Niche Adaptation.

Enterobacter cloacae complex (Ecc) species are widely distributed opportunistic pathogens mainly associated with humans and plants. In this study, the genomes of clinical isolates including E. hormaechei, E. kobei, and E. ludwigii and non-clinical isolate including E. nimipressuralis were analysed in combination with the genome of E. asburiae by using the reference strain E. cloacae subsp. cloacae ATCC 13047; the Ecc strains were tested on artificial sputum media (ASM), which mimics the host, to evaluate T6SS genes as a case study. All five Ecc strains were sequenced in our lab. Comparative genome analysis of the Ecc strains revealed that genes associated with the survival of Ecc strains, including genes of metal-requiring proteins, defence-associated genes and genes associated with general physiology, were highly conserved in the genomes. However, the genes involved in virulence and drug resistance, specifically those involved in bacterial secretion, host determination and colonization of different strains, were present in different genomic regions. For example, T6SS accessory and core components, T4SS, and multidrug resistance genes/efflux system genes seemed vital for the survival of Ecc strains in various environmental niches, such as humans and plants. Moreover, the ASM host-mimicking growth medium revealed significantly high expression of T6SS genes, including PrpC, which is a regulatory gene of the T6SS, in all tested Ecc strains compared to the control medium. The variations in T6SS gene expression in ASM vs. control showed that the ASM system represents a simple, reproducible and economical alternative to animal models for studies such as those aimed at understanding the divergence of Ecc populations. In summary, genome sequencing of clinical and environmental Ecc genomes will assist in understanding the epidemiology of Ecc strains, including the isolation, virulence characteristics, prevention and treatment of infectious disease caused by these broad-host-range niche-associated species.

Implant Sonication versus Tissue Culture for the Diagnosis of Spinal Implant Infection.

MINI: We compared the sensitivity and specificity of peri-implant tissue culture to the vortexing-sonication technique for the diagnosis of spinal implant infection (SII). Lower thresholds of sonicate fluid culture positivity showed increased sensitivity with maintained specificity. We recommend a threshold of 20 CFU/10 mL for sonicate culture positivity for the diagnosis of SII. STUDY DESIGN: This is a retrospective study comparing the diagnosis of spinal implant infection (SII) by peri-implant tissue culture to vortexing-sonication of retrieved spinal implants. OBJECTIVE: We hypothesized that vortexing-sonication would be more sensitive than peri-implant tissue culture. SUMMARY OF BACKGROUND DATA: We previously showed implant vortexing-sonication followed by culture to be more sensitive than standard peri-implant tissue culture for diagnosing of SII. In this follow-up study, we analyzed the largest sample size available in the literature to compare these two culture methods and evaluated thresholds for positivity for sonicate fluid for SII diagnosis. METHODS: We compared peri-implant tissue culture to the vortexing-sonication technique which samples bacterial biofilm on the surface of retrieved spinal implants. We evaluated different thresholds for sonicate fluid positivity and assessed the sensitivity and specificity of the two culture methods for the diagnosis of SII. RESULTS: A total of 152 patients were studied. With more than 100 colony forming units (CFU)/10 mL as a threshold for sonicate fluid culture positivity, there were 46 patients with SII. The sensitivities of peri-implant tissue and sonicate fluid culture were 65.2% and 79.6%; the specificities were 88.7% and 93.4%, respectively. With more than 50 CFU/10 mL as a threshold, there were 50 patients with SII. The sensitivities of peri-implant tissue and sonicate fluid culture were 68.0% and 76.0%; the specificities were 92.2% for both methods. Finally, with more than or equal to 20 CFU/10 mL as a threshold, there were 52 patients with SII. The sensitivities of peri-implant tissue and sonicate fluid culture were 69.2% and 82.7%; the specificities were 94.0% and 92.0%, respectively. CONCLUSION: Implant sonication followed by culture is a sensitive and specific method for the diagnosis of SII. Lower thresholds for defining sonicate fluid culture positivity allow for increased sensitivity with a minimal decrease in specificity, enhancing the clinical utility of implant sonication. LEVEL OF EVIDENCE: 4.

This is a retrospective study comparing the diagnosis of spinal implant infection (SII) by peri-implant tissue culture to vortexing­sonication of retrieved spinal implants. We hypothesized that vortexing­sonication would be more sensitive than peri-implant tissue culture. We previously showed implant vortexing­sonication followed by culture to be more sensitive than standard peri-implant tissue culture for diagnosing of SII. In this follow-up study, we analyzed the largest sample size available in the literature to compare these two culture methods and evaluated thresholds for positivity for sonicate fluid for SII diagnosis. We compared peri-implant tissue culture to the vortexing­sonication technique which samples bacterial biofilm on the surface of retrieved spinal implants. We evaluated different thresholds for sonicate fluid positivity and assessed the sensitivity and specificity of the two culture methods for the diagnosis of SII. A total of 152 patients were studied. With more than 100 colony forming units (CFU)/10 mL as a threshold for sonicate fluid culture positivity, there were 46 patients with SII. The sensitivities of peri-implant tissue and sonicate fluid culture were 65.2% and 79.6%; the specificities were 88.7% and 93.4%, respectively. With more than 50 CFU/10 mL as a threshold, there were 50 patients with SII. The sensitivities of peri-implant tissue and sonicate fluid culture were 68.0% and 76.0%; the specificities were 92.2% for both methods. Finally, with more than or equal to 20 CFU/10 mL as a threshold, there were 52 patients with SII. The sensitivities of peri-implant tissue and sonicate fluid culture were 69.2% and 82.7%; the specificities were 94.0% and 92.0%, respectively. Implant sonication followed by culture is a sensitive and specific method for the diagnosis of SII. Lower thresholds for defining sonicate fluid culture positivity allow for increased sensitivity with a minimal decrease in specificity, enhancing the clinical utility of implant sonication. Level of Evidence: 4.

Impact of Developmental Age, Necrotizing Enterocolitis Associated Stress, and Oral Therapeutic Intervention on Mucus Barrier Properties.

Necrotizing enterocolitis (NEC) is a devastating gastrointestinal disease of incompletely understood pathophysiology predominantly affecting premature infants. While NEC is associated with microbial invasion of intestinal tissues, and mucus modulates interactions between microbes and underlying tissues, variations in mucus barrier properties with NEC-associated risk factors have not been investigated. This study explored differences in mucus composition (total protein, DNA, mucin content, sialic acid, and immunoregulatory proteins), as well as structural and transport properties, assessed by tracking of particles and bacteria (E. coli and E. cloacae) with developmental age and exposure to NEC stressors in Sprague Dawley rats. Early developmental age (5 day old) was characterized by a more permeable mucus layer relative to 21 day old pups, suggesting immaturity may contribute to exposure of the epithelium to microbes. Exposure to NEC stressors was associated with reduced mucus permeability, which may aid in survival. Feeding with breastmilk as opposed to formula reduces incidence of NEC. Thus, NEC-stressed (N-S) rat pups were orally dosed with breastmilk components lysozyme (N-S-LYS) or docosahexaenoic acid (N-S-DHA). N-S-LYS and N-S-DHA pups had a less permeable mucus barrier relative to N-S pups, which suggests the potential of these factors to strengthen the mucus barrier and thus protect against disease.

Transcriptome analysis and immune-related genes expression reveals the immune responses of Macrobrachium rosenbergii infected by Enterobacter cloacae.

Macrobrachium rosenbergii is an important cultural species in China and other Southeast Asian countries. However, Enterobacter cloacae infection has caused a great economic loss in M. rosenbergii culture industry. The immune responses of M. rosenbergii to the E. cloacae infection is not fully characterized. To investigate the immune response of M. rosenbergii against E. cloacae, we performed transcriptome analysis of the M. rosenbergii hepatopancreas with and without E. cloacae infection using RNA-seq. After assembly and annotation, 29,731 high quality unigenes were obtained from RNA-seq data. Differential expression analysis revealed the existence of 2498 significantly differently expressed genes (DEGs) at 12 h post infection, with 1365 up-regulated and 1133 down-regulated genes. Among these DEGs, some well-known immune-related genes were up-regulated significantly, including C-type lectin 1, lectin 3, anti-lipopolysaccharide factor 2, Cu/Zn superoxide dismutase and heat shock protein 70. GO analysis demonstrated 24 biological process subcategories, 14 cellular component subcategories, and 12 molecular function subcategories that were enriched among these DEGs, and some DEGs were clustered into immune related subcategories such as immune system process, response to stimulus, biological adhesion, and antioxidant activity. These DEGs were enriched into 216 KEGG pathways including a core set of immune correlated pathways notably in phagosome and lysosome. In addition, 5 up-regulated and 5 down-regulated immune-related DEGs were selected for further validation by quantitative real-time PCR and the results showed consistence with the RNA-seq data. Additionally, the expression level of six selected immune-related genes (ALF2, CLEC1, LEC3, hemocyanin1, HSP70 and SOD) based on the transcriptomic data were monitored at different point of time in hepatopancreas, gill, hemolymph and intestine. Results revealed these immune-related genes were significantly up-regulated in different tissues from 6 to 24 h after E. cloacae infection. Overall, these results provided valuable information for further studying the immune response of M. rosenbergii against E. cloacae infection.

Improvement of rice plant productivity by native Cr(VI) reducing and plant growth promoting soil bacteria Enterobacter cloacae.

Rapid industrialization and anthropogenic activities have produced huge amount of noxious Cr(VI), which accumulate in the soil for longer period. As a consequence, that decreases rice plant productivity in contiguous agricultural field of Sukinda mining area, Odisha. Thus, the high Cr(VI) resistant native bacterial strain CTWI-06 was selected for the study, which depicted resistance to 3500 ppm of Cr(VI) and wide array of other metals. Under optimized condition, the multi-metal resistant bacteria reduced 94% Cr(VI) within 92 h and Cr(VI) reduction was confirmed by FTIR and XRD analysis. Plant growth promoting traits like N2 fixation; phosphate (146.87 ppm), potassium (12.55 ppm) and Zn solubilization; ammonification; IAA production (114 µg mL-1) and suppression of fungal phytopathogens such as Rhizoctonia solani (ITCC 2060) and Phytium debaryanum (ITCC 5488) were also recorded. The bacterial strain was identified as Enterobacter cloacae CTWI-06 by 16S rDNA sequence (Accession No. MG757378). It significantly improved growth traits as well as productivity of Mahalakshmi rice variety in pot culture. Thus, the potential Cr(VI) reducing and PGPB strain may be utilized for long term bioremediation of Cr(VI) in chromium contaminated soil and to maintain soil fertility.

RETRACTED: Antibacterial and antibiofilm activity of ursolic acid against carbapenem-resistant Enterobacter cloacae.

This article has been retracted: please see Elsevier Policy on Article Withdrawal (https://www.elsevier.com/about/our-business/policies/article-withdrawal). This article has been retracted at the request ofthe corresponding author and the editor-in-chief. The authors have notified the journal that Fig. 3E and 3F in this article was a duplicate image of Fig. 3C of the article published by Weidong Qian, Miao Liu, Yuting Fu, Ting Wang, Jianing Zhang, Min Yang, Zhaohuan Sun, Xiang Li, and Yongdong Li in Foodborne Pathogens and Disease 129 (2020) 528-534 https://www.liebertpub.com/doi/10.1089/fpd.2019.2751 and requested for retraction. The authors have subsequently admitted that Figs. 3A, 3D and 4B in this article were duplicated from Figs. 3A, 5A and 7B of the article published by Weidong Qian, Yuting Fu, Miao Liu, Ting Wang, Jianing Zhang, Min Yang, Zhaohuan Sun, Xiang Li and Yongdong Li in Antibiotics 8 (2019) 220 https://www.mdpi.com/2079-6382/8/4/220, which has been retracted https://www.mdpi.com/2079-6382/9/6/322. The authors have claimed that there were errors in the preparation of figures for three papers which had been submitted to multiple journals from August to October 2019. The authors have provided replacement images for Figs. 3 and 4 of this article, but these data did not resolve the concerns that call into question the reliability of the study findings. In light of these issues, the journal has decided to retract the article. The authors assert the findings, that ursolic acid exerted antibacterial activity against carbapenem-resistant Enterobacter cloacae, are nevertheless reliable.

Motility effects biofilm formation in Pseudomonas aeruginosa and Enterobacter cloacae.

Chronic infections caused by gram negative bacteria are the mains reasons to have morbidity and death in patients, despite using high doses of antibiotics applied to cure diseases producing by them. This study was designed to identify the role of flagella in biofilm formation Ten pure strains were collected from our lab. Morphological variation and motility assays led us to study two strains in detail. They were characterized biochemically, physiologically and genetically. Biofilm formation analysis was performed using test tube assay, congo red assay and liquid-interface coverslip assay. In order to disrupt flagella of studied strains, blending was induced for 5, 10 and 15 minutes followed by centrifugation and observing motility using motility test. Biofilm quantification of wild type (parental) and blended strains was done using test tube and liquid interface coverslip assays. 16S rRNA sequencing identified strains as Pseudomonas aeroginosa and Enterobacter cloacae. Significant biofilm formation (p>0.05) by was observed after 72 and 18 hours using test tube and liquid-interface coverslip assays respectively. Flagellar disruption showed that 15 minutes blending caused significant reduction in both strains, hence demonstrated that flagellar mediated motility could be a potent strategy to stabilize aggregate and invest resources for biofilm formation in P. aeruginosa and E. cloacae.

Antiobesity Effects of Lactobacillus plantarum LMT1-48 Accompanied by Inhibition of Enterobacter cloacae in the Intestine of Diet-Induced Obese Mice.

The gut microbiota is the most important environmental factor that plays a role in inducing obesity. The gram-negative bacteria, Enterobacter cloacae strains, recently identified in obese mice are considered to be pathogenic bacteria in the gut. Probiotics are important members of the gut microbiota and exert beneficial effects, including inhibiting the growth of potential pathogenic bacteria. Therefore, we isolated a total of 230 lactic acid bacteria from traditional, Korean fermented foods and fecal samples from newborn infants, including Lactobacillus plantarum LMT1-48, which exhibited maximal antimicrobial activity against E. cloacae. We next investigated the functional antiobesity effects of L. plantarum LMT1-48 in an E. cloacae-induced high-fat diet (HFD)-fed animal obesity model. To this end, the L. plantarum LMT1-48 showed antiobesity effects, including body weight loss and reduction of abdominal fat volume, which was accompanied by a decrease in leptin and total cholesterol levels in E. cloacae-induced HFD-fed mice. Notably, gut microbiota diversity also increased after long-term ingestion of L. plantarum LMT1-48, resulting in amelioration of obesity in E. cloacae-induced HFD-fed mice. Accordingly, results suggest that dietary intake of L. plantarum LMT1-48 protects against the onset of E. cloacae-induced obesity.

Connecting single-cell properties to collective behavior in multiple wild isolates of the Enterobacter cloacae complex.

Some strains of motile bacteria self-organize to form spatial patterns of high and low cell density over length scales that can be observed by eye. One such collective behavior is the formation in semisolid agar media of a high cell density swarm band. We isolated 7 wild strains of the Enterobacter cloacae complex capable of forming this band and found its propagation speed can vary 2.5 fold across strains. To connect such variability in collective motility to strain properties, each strain's single-cell motility and exponential growth rates were measured. The band speed did not significantly correlate with any individual strain property; however, a multilinear analysis revealed that the band speed was set by a combination of the run speed and tumbling frequency. Comparison of variability in closely-related wild isolates has the potential to reveal how changes in single-cell properties influence the collective behavior of populations.

Shifts along the parasite-mutualist continuum are opposed by fundamental trade-offs.

Theory suggests that symbionts can readily evolve more parasitic or mutualistic strategies with respect to hosts. However, many symbionts have stable interactions with hosts that improve nutrient assimilation or confer protection from pathogens. We explored the potential for evolution of increased parasitism or decreased parasitism and mutualism in a natural gut symbiosis between larvae of Plutella xylostella and the microbe Enterobacter cloacae. We focused on interactions with the pathogen, Bacillus thuringiensis: selecting for parasitism in terms of facilitating pathogen infection, or increased mutualism in terms of host protection. Selection for parasitism led to symbionts increasing pathogen-induced mortality but reduced their competitive ability with pathogens and their in vitro growth rates. Symbionts did not evolve to confer protection from pathogens. However, several lineages evolved reduced parasitism, primarily in terms of moderating impacts on host growth, potentially because prudence pays dividends through increased host size. Overall, the evolution of increased parasitism was achievable but was opposed by trade-offs likely to reduce fitness. The evolution of protection may not have occurred because suppressing growth of B. thuringiensis in the gut might provide only weak protection or because evolution towards protective interactions was opposed by the loss of competitive fitness in symbionts.

Specific interactions of rotavirus HAL1166 with Enterobacter cloacae SENG-6 and their contribution on rotavirus HAL1166 removal.

Contribution of specific interactions between human enteric viruses and wastewater suspended solids on human enteric virus removal by microfiltration was studied. A cross-flow microfiltration system was used with rotavirus HAL1166 and Enterobacter cloacae SENG-6 as the model virus and wastewater suspended solid. Cleavage of rotavirus HAL1166 protein VP4 by trypsin produces the VP8* subunit, which specifically interacts with histo-blood group antigen (HBGA). In the presence of Enterobacter cloacae SENG-6, the trypsin-treated rotavirus concentration reduced with time (R2 > 0.6) compared to the reduction of non-trypsin treated rotavirus. Calculation of the gel/cake layer deposited on the membrane, consisting of Enterobacter cloacae SENG-6 and either trypsin-treated or non-trypsin treated rotavirus HAL1166, revealed that the microflocs consisting of trypsin-treated rotavirus and Enterobacter cloacae SENG-6 have lower porosity and permeability, displaying higher resistance to virus passage through the membrane. The results provide evidence that specific wastewater suspended solids-human enteric virus interaction can contribute to increasing the removal of human enteric viruses by microfiltration.

Surfing Motility: a Conserved yet Diverse Adaptation among Motile Bacteria.

Bacterial rapid surfing motility is a novel surface adaptation of Pseudomonas aeruginosa in the presence of the glycoprotein mucin. Here, we show that other Gram-negative motile bacterial species, including Escherichia coli, Salmonella enterica, Vibrio harveyi, Enterobacter cloacae, and Proteus mirabilis, also exhibit the physical characteristics of surfing on the surface of agar plates containing 0.4% mucin, where surfing motility was generally more rapid and less dependent on medium viscosity than was swimming motility. As previously observed in Pseudomonas aeruginosa, all surfing species exhibited some level of broad-spectrum adaptive resistance, although the antibiotics to which they demonstrated surfing-mediated resistance differed. Surfing motility in P. aeruginosa was found to be dependent on the quorum-sensing systems of this organism; however, this aspect was not conserved in other tested bacterial species, including V. harveyi and S. enterica, as demonstrated by assaying specific quorum-sensing mutants. Thus, rapid surfing motility is a complex surface growth adaptation that is conserved in several motile bacteria, involves flagella, and leads to diverse broad-spectrum antibiotic resistance, but it is distinct in terms of dependence on quorum sensing.IMPORTANCE This study showed for the first time that surfing motility, a novel form of surface motility first discovered in Pseudomonas aeruginosa under artificial cystic fibrosis conditions, including the presence of high mucin content, is conserved in other motile bacterial species known to be mucosa-associated, including Escherichia coli, Salmonella enterica, and Proteus mirabilis Here, we demonstrated that key characteristics of surfing, including the ability to adapt to various viscous environments and multidrug adaptive resistance, are also conserved. Using mutagenesis assays, we also identified the importance of all three known quorum-sensing systems, Las, Rhl, and Pqs, in P. aeruginosa in regulating surfing motility, and we also observed a conserved dependence of surfing on flagella in certain species.

Enterobacter cloacae administration induces hepatic damage and subcutaneous fat accumulation in high-fat diet fed mice.

Accumulating evidence indicates that gut microbiota plays a significant role in obesity, insulin resistance and associated liver disorders. Family Enterobacteriaceae and especially Enterobacter cloacae strain B29 have been previously linked to obesity and hepatic damage. The underlying mechanisms, however, remain unclear. Therefore, we comprehensively examined the effects of E. cloacae subsp. cloacae (ATCC® 13047™) administration on host metabolism of mice fed with high-fat diet (HFD). C57BL/6N mice were randomly divided into HFD control, chow control, and E. cloacae treatment groups. The E. cloacae treatment group received live bacterial cells in PBS intragastrically twice a week, every other week for 13 weeks. Both control groups received PBS intragastrically. After the 13-week treatment period, the mice were sacrificed for gene and protein expression and functional analyses. Our results show that E. cloacae administration increased subcutaneous fat mass and the relative proportion of hypertrophic adipocytes. Both subcutaneous and visceral fat had signs of decreased insulin signaling and elevated lipolysis that was reflected in higher serum glycerol levels. In addition, E. cloacae -treated mice had significantly higher hepatic AST and AST/ALT ratio, and their liver histology indicated fibrosis, demonstrating that E. cloacae subsp. cloacae administration promotes hepatic damage in HFD fed mice.