Exploring the protist microbiome: the diversity of bacterial communities associated with Arcella spp. (Tubulina: Amoebozoa)

© The Author(s), 2022. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Gomaa, F., Utter, D. R., Loo, W., Lahr, D. J. G., & Cavanaugh, C. M. Exploring the protist microbiome: the diversity of bacterial communities associated with Arcella spp. (Tubulina: Amoebozoa). European Journal of Protistology, 82, (2022): 125861, https://doi.org/10.1016/j.ejop.2021.125861.Research on protist-bacteria interactions is increasingly relevant as these associations are now known to play important roles in ecosystem and human health. Free-living amoebae are abundant in all environments and are frequent hosts for bacterial endosymbionts including pathogenic bacteria. However, to date, only a small fraction of these symbionts have been identified, while the structure and composition of the total symbiotic bacterial communities still remains largely unknown. Here, we use the testate amoeba Arcella spp. as model organisms to investigate the specificity and diversity of Arcella-associated microbial communities. High-throughput amplicon sequencing from the V4 region of the 16S rRNA gene revealed high diversity in the bacterial communities associated with the wild Arcella spp. To investigate the specificity of the associated bacterial community with greater precision, we investigated the bacterial communities of two lab-cultured Arcella species, A. hemispherica and A. intermedia, grown in two different media types. Our results suggest that Arcella-bacteria associations are species-specific, and that the associated bacterial community of lab-cultured Arcella spp. remains distinct from that of the surrounding media. Further, each host Arcella species could be distinguished based on its bacterial composition. Our findings provide insight into the understanding of eukaryotic-bacterial symbiosis.This project was funded by National Science Foundation Postdoctoral Research Fellowship in Biology to F. Gomaa, Grant Number: PRFB1611514. Support was provided to D.R.U. from the National Science Foundation Graduate Research Fellowship Program under Grant No. DGE1745303 to D.R.U and by Harvard University’s Department of Organismic and Evolutionary Biology program

Toward establishing model organisms for marine protists : Successful transfection protocols for Parabodo caudatus (Kinetoplastida: Excavata)

Author Posting. © The Author(s), 2017. This is the author's version of the work. It is posted here under a nonexclusive, irrevocable, paid-up, worldwide license granted to WHOI. It is made available for personal use, not for redistribution. The definitive version was published in Environmental Microbiology 19 (2017): 3487-3499, doi:10.1111/1462-2920.13830.We developed protocols for, and demonstrated successful transfection of, the free-living kinetoplastid flagellate Parabodo caudatus with three plasmids carrying a fluorescence reporter gene (pEF-GFP with the EF1 alpha promoter, pUB-GFP with Ubiquitin C promoter, and pEYFP37 Mitotrap with CMV promoter). We evaluated three electroporation approaches: 1) a square-wave electroporator designed for eukaryotes, 2) a novel microfluidic transfection system employing hydrodynamically-controlled electric field waveforms, and 3) a traditional exponential decay electroporator. We found the microfluidic device provides a simple and efficient platform to quickly test a wide range of electric field parameters to find the optimal set of conditions for electroporation of target species. It also allows for processing large sample volumes (> 10 ml) within minutes, increasing throughput 100 times over cuvettes. Fluorescence signal from the reporter gene was detected a few hours after transfection and persisted for 3 days in cells transformed by pEF-GFP and pUB-GFP plasmids and for at least 5 days post-transfection for cells transformed with pEYFP-Mitotrap. Expression of the reporter genes (GFP and YFP) was also confirmed using reverse transcription-PCR (RT-PCR). This work opens the door for further efforts with this taxon and close relatives toward establishing model systems for genome editing.This project was funded by the Gordon and Betty Moore Foundation through Grant GBMF4963 to V. Edgcomb, P. Girguis, and C. Buie

Vancomycin MIC Distribution among Methicillin-Resistant Staphylococcus Aureus. Is Reduced Vancomycin Susceptibility Related To MIC Creep?

AIM: To determine the distribution of vancomycin MIC and the frequency of S. aureus strains with reduced vancomycin susceptibility among Methicillin-Resistant Staphylococcus aureus (MRSA) isolates. METHODS: MRSA isolates (n = 100) were tested for reduced susceptibility to vancomycin using  MIC broth microdilution method (BMD), vancomycin screening agar with different vancomycin concentrations with and without casein, and Vitek 2 system. RESULTS: BMD detected (22%) vancomycin-intermediate S. aureus (VISA) and (78%) vancomycin-susceptible S. aureus (VSSA) but couldn’t detect nine (Heterogeneous VISA) (hVISA)   isolates (9%) with MIC ≤ 2 µg/ml that grew on screening agar 4 µg/ml or 6 µg/ml. Adding casein to vancomycin screening agar increased detection rate of VISA by 4.5%.  Screening agar with 6 µg/ml vancomycin overall detection rate for VISA was 95.45%. Probable ‘pre-hVISA’isolates (17%) showed growth on vancomycin screening agar 2 µg/ml with casein. Vitek 2 system failed to detect any VISA isolates. CONCLUSION: Vancomycin screening agar; 2 µg/ml and (4 and 6 µg/ml) were able to detect; probable “pre hVISA and (hVISA and VISA) isolates respectively based on their BMD MIC values. Decreased vancomycin susceptibility in MRSA isolates might be related to MIC creep. Analysis of vancomycin MIC values over longer periods is recommended to further study this phenomenon and its impact on vancomycin treatment failure.   ABSTRACT Aims: Determine the distribution of vancomycin MIC and the frequency of S. aureus strains with reduced vancomycin susceptibility among MRSA isolates.  Methods:  MRSA isolates (n =100) were tested for reduced susceptibility to vancomycin using  MIC broth microdilution method(BMD), vancomycin screening agar with different vancomycin concentrations with and without casein, and Vitek 2 system.  Results: BMD detected (22%) vancomycin intermediate S. aureus(VISA) and (78%) vancomycin susceptible S. aureus(VSSA) but failed to detect nine (Heterogeneous VISA) (hVISA)  isolates (9%) with MIC ≤2ug/ml that grew on screening agar 4ug/ml or 6 ug/ml. Adding casein to vancomycin screening agar increased detection rate of VISA by 4.5%.  Screening agar with 6 ug/ml vancomycin over all detection rate for VISA was 95.45%. Probable ‘pre-hVISA’isolates (17%) showed growth on vancomycin screening agar 2µg/ml with casein. Vitek 2 system failed to detect any VISA isolates. Conclusion: vancomycin screening agar; 2 µg/ml and (4 and 6 µg/ml) were able to detect; probable “pre hVISA and (hVISA and VISA) isolates respectively based on their BMD MIC values. Decreased vancomycin susceptibility in MRSA isolates might be related to MIC creep. Analysis of vancomycin MIC values over longer periods of time is recommended to further study this phenomenon and its impact on vancomycin treatment failure.   &nbsp

Characterization and genomic analysis of the lytic bacteriophage vB_EclM_HK6 as a potential approach to biocontrol the spread of Enterobacter cloacae contaminating food

Background: Increased prevalence of Enterobacter cloacae within food products underscores food as an underexplored reservoir for antibiotic resistance, thus requiring particular intervention. Bacteriophages have been explored as a promising approach for controlling bacterial growth in different matrices. Moreover, their specific interaction and self-replication, put them apart from traditional methods for controlling bacteria in different matrices. Methods: Sixteen Enterobacter cloacae strains were recovered from raw chicken. These strains were used to isolate bacteriophages using enrichment protocol. The broad-spectrum bacteriophage was evaluated in terms of thermal, pH, shearing stress and storge. Moreover, its infection kinetics, in vitro antibacterial activity, cytotoxicity were also assessed. Genomic sequencing was performed to exclude any potential virulence or resistance genes. Finally, the capability of the isolated phages to control bacterial growth in different chicken samples was assessed alone and in combination with sodium nitrite. Results: The lytic bacteriophage vB_EclM_HK6 was isolated and showed the broadest spectrum being able to infect 8/16 E. cloacae strains with a lytic activity against its host strain, E. cloacae EC21, as low as MOI of 10–6. The phage displays a latent period of 10 min and burst size of 115 ± 44 and resistance frequency of 5.7 × 10–4 ± 3.0 × 10–4. Stability assessment revealed a thermal tolerance up to 60 ˚C, wide range pH stability (3–10) and the ability to withstand shearing stress up to 250 rpm. HK6 shows no cytotoxicity against oral epithelial cells up to 1012 PFU/ml. Genomic analysis revealed a Strabovirus with total size of 177,845 bp that is free from known resistance and virulence genes. Finally, HK6 pretreatment of raw chicken, chicken nuggets and ready-made cheese salad shows a reduced bacterial count up to 4.6, 2.96 and 2.81 log-units, respectively. Moreover, combing HK6 with sodium nitrite further improved the antibacterial activity in both raw chicken and chicken nuggets without significant enhancement in case of cheese salad. Conclusion: Enterobacter bacteriophage vB_EclM_HK6 presents a safe and effective approach for controlling E. cloacae contaminating stored chicken food samples. Moreover, they could be combined with a reduced concentrations of sodium nitrite to improve the killing capacity

Novel 1,5-diaryl pyrazole-3-carboxamides as selective COX-2/sEH inhibitors with analgesic, anti-inflammatory, and lower cardiotoxicity effects

Funding Information: The authors extend their appreciation to the Deanship of Scientific Research at Jouf University for funding this work through research grant number (DSR2020-04-421 )Peer reviewedPostprin

Essential Facts on the History of Hyperthermia and their Connections with Electromedicine

The term hyperthermia is a combination of two Greek words: HYPER (rise) and THERME (heat) and refers to the increasing of body temperature or selected tissues in order to achieve a precise therapeutic effect. This paper reviews the development of thermotherapy by describing the most important moments in its history. For decades, the development of hyperthermia ran parallel with the development of cancer treatment and had numerous connections with electromedicine. Throughout its history, hyperthermia evoked a number of hopes, brought spectacular successes, but also was the subject of many disappointments

Discovery of novel oxazole-based macrocycles as anti-coronaviral agents targeting SARS-CoV-2 main protease

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