Enrichment Culture Research Articles

Impact of TiO2, ZnO, and Ag nanoparticles on anammox activity in enriched river Nile sediment cultures: unveiling differential effects and environmental implications.

The increasing use of nanoparticles (NPs) necessitates investigation of their impact on wastewater treatment processes, particularly anammox, a critical biological nitrogen removal pathway. This study explored the effects of short-term exposure to TiO2, ZnO, and Ag-NPs on anammox activity in enriched cultures derived from River Nile sediments. Anammox bacteria were identified and enriched, with activity confirmed through 16S rRNA and hydrazine oxidoreductase (hzo) gene amplification and sequencing. Activity assays demonstrated efficient ammonium removal by the enriched culture. Subsequently, the impact of different sized and concentrated NPs on anammox activity was assessed. XRD analysis confirmed NP behavior within the microcosms: TiO2 transformed, ZnO partially dissolved, and Ag remained ionic. hzo gene expression served as a biomarker for anammox bacterial activity. Interestingly, 100nm TiO2-NPs up-regulated hzo expression, potentially indicating a non-inhibitory transformed phase. Conversely, ZnO and Ag-NPs across all sizes and concentrations significantly down-regulated hzo expression, suggesting detrimental effects. Ag-NPs amended microcosms showed a significant reduction (79%) in hzo gene expression and a detrimental effect on bacterial populations. Overall, anammox activity mirrored hzo expression patterns, with TiO2 (21 and 25nm, respectively) exhibiting the least inhibition, followed by ZnO and Ag-NPs. This study highlights the differential effects of NPs on anammox, with the order of impact being Ag > ZnO > TiO2. These findings provide valuable insights into the potential environmental risks of NPs on anammox-mediated nitrogen cycling in freshwater ecosystems.

Potential role of alginate in marine bacteria-yeast interactions.

The ability of microorganisms to decompose brown algae has attracted attention. This study aims to clarify the characteristics of marine microbial communities in which prokaryotic and eukaryotic microorganisms interact via the metabolism of brown algae carbohydrates. Amplicon-based microbiome analysis revealed the predominance of the genera Marinomonas and Vibrio in seawater and seaweed samples mixed with alginate and mannitol, which are the primary carbohydrates in brown algae. Three Vibrio species and Candida intermedia were isolated via alginate enrichment culture. Although C. intermedia did not utilize alginate as a nutrient source, the yeast grew in the spent alginate medium in which Vibrio algivorus had been cultured. Coculture with C. intermedia and the Vibrio isolates, especially V. algivorus, also enhanced the growth of the yeast on alginate. These results suggested that C. intermedia grew because of the supply of nutrients via alginate metabolism by Vibrio species. In the coculture medium, the amount of phosphatidylserine increased in the early phase but decreased with the growth of C. intermedia, indicating that phosphatidylserine secreted by Vibrio is involved in the putative mutualistic interaction. We examined whether such interaction is applicable to the production of useful substances and succeeded in lipid production by oleaginous marine yeast Yarrowia lipolytica through coculture with V. algivorus. Our study suggested the potential of mutualistic interaction via degradation of alginate by marine Vibrio for production of industrially useful substances in yeast cells.IMPORTANCEIn this study, we analyzed the microbiome of seawater and seaweed in the presence of brown algae carbohydrates and reconstructed the putative mutualistic relationship of marine Vibrio and Candida intermedia mediated by metabolism of brown algae in the ocean.

Verrucomicrobia of the Family Chthoniobacteraceae Participate in Xylan Degradation in Boreal Peat Soils

The phylum Verrucomicrobiota is one of the main groups of soil prokaryotes, which remains poorly represented by cultivated organisms. The major recognized role of Verrucomicrobiota in soils is the degradation of plant-derived organic matter. These bacteria are particularly abundant in peatlands, where xylan-type hemicelluloses represent one of the most actively decomposed peat constituents. The aim of this work was to characterize the microorganisms capable of hydrolyzing xylan under the anoxic conditions typical of peatland soils. The laboratory incubation of peat samples with xylan resulted in the pronounced enrichment of several phylotypes affiliated with the Verrucomicrobiota, Firmicutes, and Alphaproteobacteria. Sequencing of the metagenome of the enrichment culture allowed us to recover high-quality metagenome-assembled genomes (MAGs) assigned to the genera Caproiciproducens, Clostridium, Bacillus (Firmicutes), and Rhizomicrobium (Alphaproteobacteria), Cellulomonas (Actinobacteriota) and the uncultured genus-level lineage of the family Chthoniobacteraceae (Verrucomicrobiota). The latter bacterium, designated “Candidatus Chthoniomicrobium xylanophilum” SH-KS-3, dominated in the metagenome and its MAG was assembled as a complete closed chromosome. An analysis of the SH-KS-3 genome revealed potential endo-1,4-beta-xylanases, as well as xylan beta-1,4-xylosidases and other enzymes involved in xylan utilization. A genome analysis revealed the absence of aerobic respiration and predicted chemoheterotrophic metabolism with the capacity to utilize various carbohydrates, including cellulose, and to perform fermentation or nitrate reduction. An analysis of other MAGs suggested that Clostridium and Rhizomicrobium could play the role of primary xylan degraders while other community members probably took advantage of the availability of xylo-oligosaccharides and xylose or utilized low molecular weight organics.

The Effects of Model Insoluble Copper Compounds in a Sedimentary Environment on Denitrifying Anaerobic Methane Oxidation (DAMO) Enrichment

The contribution of denitrifying anaerobic methane oxidation (DAMO) as a methane sink across different habitats, especially those affected by anthropogenic activities, remains unclear. Mining and industrial and domestic use of metals/metal-containing compounds can all cause metal contamination in freshwater ecosystems. Precipitation of metal ions often limits their toxicity to local microorganisms, yet microbial activity may also cause the redissolution of various precipitates. In contrast to most other studies that apply soluble metal compounds, this study investigated the responses of enriched DAMO culture to model insoluble copper compounds, malachite and covellite, in simulated sedimentary environments. Copper ≤ 0.22 µm from covellite appeared to cause immediate inhibition in 10 h. Long-term tests (54 days) showed that apparent methane consumption was less impacted by various levels of malachite and covellite than soluble copper. However, the medium-/high-level malachite and covellite caused a 46.6–77.4% decline in denitrification and also induced significant death of the representative DAMO microorganisms. Some enriched species, such as Methylobacter tundripaludum, may have conducted DAMO or they may have oxidized methane aerobically using oxygen released by DAMO bacteria. Quantitative polymerase chain reaction analysis suggests that Candidatus Methanoperedens spp. were less affected by covellite as compared to malachite while Candidatus Methylomirabilis spp. responded similarly to the two compounds. Under the stress induced by copper, DAMO archaea, Planctomycetes spp. or Phenylobacterium spp. synthesized PHA/PHB-like compounds, rendering incomplete methane oxidation. Overall, the findings suggest that while DAMO activity may persist in ecosystems previously exposed to copper pollution, long-term methane abatement capability may be impaired due to a shift of the microbial community or the inhibition of representative DAMO microorganisms.

Acid mine drainage bioremediation using bacteria enriched from the confluence zone between its flow and treated sewage

AbstractSediments from the confluence zone between an acid mine drainage stream and a water stream receiving the effluent from a municipal wastewater treatment plant were inoculated in a Postgate B medium base having methanol, glycerol, or ethanol as carbon source/electron donor, over a pH range between 2.00 and 6.00, aiming to obtain native cultures enriched in acidophilic/acid-tolerant sulfate-reducing bacteria. The most effective sulfate reduction observed in acidic conditions was in the enrichment cultures with methanol. Thus, the microbial consortium enriched in these conditions was further used in acid mine drainage bioremediation experiments at an initial pH of 4.50 and using different doses of nutrients medium base and methanol as carbon source/electron donor. The most promising results, with more than 99% removal of metals, were obtained in the mixture of 20% Postgate B medium base plus 80% acid mine drainage (v/v), which corresponds to 0.115% basal salts and 0.02% yeast extract (w/v). Metataxonomic analysis based on 16S rRNA gene sequences showed the presence of Desulfosporosinus spp. in all enrichment cultures, with the highest relative abundance in cultures at pH 4.00 with methanol. Furthermore, metataxonomic analysis in the acid mine drainage remediation tests revealed the presence of Desulfosporosinus spp. in all tested conditions. In addition, the study also revealed Clostridium members in all tests. Plus, their relative abundances were related to the dose of nutrient medium base and in balance with the abundances of Desulfosporosinus spp.

Defluorination of various perfluoro alkyl acids and selected PFOA and PFOS monomers by Acidimicrobium sp. Strain A6 enrichment cultures

Per- and polyfluoroalkyl substances (PFAS) have emerged as a diverse class of environmental pollutants, garnering increasing attention due to their various structural types and potential ecological impacts. The impact of select PFAS on environmental microorganisms and the potential for microbial degradation of certain PFAS are timely research topics. In this study, we conducted a series of batch incubation to investigate the effects of C4-C10 perfluoroalkyl carboxylic acids (PFCAs) and perfluorosulfonic acids (PFSAs), as well as linear and branched perfluorooctanoic acid (PFOA) and perfluorooctanesulfonic acid (PFOS) monomers, on the Feammox reaction and Acidimicrobium sp. A6 (A6), a microbe known to degrade PFOA and PFOS. We explored the defluorination ability of A6 cultures with these PFAS, evaluating their response to varying chemical structures. While A6 cultures demonstrated the ability to degrade a wide range of PFAAs (11.5–56.9 % reduction over 120 days), challenges were noted with specific compounds like PFPeA and double-branched PFCAs and PFSAs, which also showed reduced ammonium removal. Additionally, exposure to the selected PFAS resulted in notable shifts in the microbial community within the A6 enrichment cultures, indicating a selective pressure that benefits certain strains (e.g., increased percentages of Acidimicrobium, Paraburkholderia, and Desulfosporosinus in several PFCA, PFSA and PFOA/PFOS monomers enriched cultures). These insights contribute to our understanding of microbial-PFAS interactions and are instrumental in developing bioremediation strategies for PFAS-impacted environments.

Genome sequence of Nitrosopumilus adriaticus CCS1 assembled from an ammonia-oxidizing enrichment culture.

We report the metagenome-assembled genome of an ammonia-oxidizing archaeon that is closely related to Nitrosopumilus adriaticus NF5 but shows distinct genomic features compared to strain NF5.

Analysis of a persistent outbreak with vancomycin resistant Enterococcus faecium (VREfm) revealed the need for an adapted diagnostic algorithm

Our setting was challenged with an outbreak of different vancomycin resistant Enterococcus faecium (VREfm) including vanA and/or vanB containing isolates. Remarkably, it was observed that screening by use of a vanA and vanB real-time PCR on overnight enriched specimens from time to time tested positive for VanB with very low Ct-values, whereas VREfm-specific enrichment cultures remained negative. Here, we describe the analysis of the diagnostic results leading to adaptation of the diagnostic algorithm. Per specimen of each patient, results of the vanA and vanB screening PCR and of the VREfm-specific culture (Brilliance VRE) were collected and combined with genotyping data of the identified VREfm isolates. During the outbreak, a second VREfm-specific culture medium (CHROMagar VRE) was introduced, and results were compared to the results obtained with Brilliance VRE agar. Thirty-five patients were identified as VREfm-carrier, in which four different strains were identified comprising vanA (STnew-CT7088) and/or vanB (ST80-CT1065, ST117-CT7117 and ST117-CT7118). Complementing results of PCR, culture and genotyping revealed that culture with Brilliance VRE agar was inadequate for detection of the vanB ST117 isolates identified, irrespective of vancomycin MIC values. In contrast, CHROMagar VRE was able to correctly detect these vanB ST117 isolates and other tested isolates. Our data showed that the vanB ST117 containing isolates were inadequately detected by the VREfm-specific culture media, possibly contributing to unnoticed spread of VREfm. For this reason, CHROMagar VRE was evaluated during the outbreak and subsequently implemented in routine diagnostics, replacing Brilliance VRE agar.

P40 A challenging case of new systemic lupus erythematosus

Abstract Introduction The diagnosis of systemic lupus erythematosus (SLE) and its distinction from systemic infection can be challenging. We present a case of new systemic lupus alongside endocarditis, which illustrates this problem. Case description A previously well, 35-year-old male tarmacer of mixed ethnicity, presented with a three months’ history of general malaise, arthralgia and weight loss of 20kg. Two weeks prior to admission he developed progressive confusion, headaches and imbalance. Clinical assessment revealed a pyrexial patient in delirium without focal neurological signs, a faint maculopapular facial rash, symmetrical synovitis, very few nailfold infarcts, a systolic and diastolic murmur and sterile haemato-proteinuria. No history of IV drug abuse. Investigations showed elevated ESR 61, CRP 217, WBC 11.70 and procalcitonin 7.0. He had AKI stage 2 (eGFR 35 with a urinary protein/creat ratio 162; homogenous ANA (1:1280) with positive dsDNA and crithidia-luciliae binding. C3 and C4 suppressed (0.59 and 0.13, respectively); One blood culture prior to broad spectrum antibiotics was negative. Assuming the possibility of Libman-Sacks endocarditis, the attending acute stroke physician initially also treated with pulsed IV steroids, as well as antibiotics. Due to the delirium, key diagnostic tests were technically difficult, although a transthoracic ECHO showed moderately severe aortic and mitral regurgitation that were not felt to be typical for infective endocarditis by the attending cardiologist. The ITU team offered sedation and controlled ventilation to enable all necessary investigations: trans-oesophageal ECHO showed vegetations of aortic and mitral valve leaflets with perforations of single leaflets, resulting in severe acute aortic and mitral regurgitation; a renal biopsy showed class 3 lupus nephritis and an MRI/A Brain showed widespread cerebral foci of restricted diffusion imaging. Consultation with the regional cardiothoracic centre advised urgent transfer for double metal valve replacement, performed on Day 8 post admission. Surgical inspection confirmed an aortic root abscess, and enrichment culture of the native valve subsequently grew Staph. aureus. In addition to IV antibiotics for infective endocarditis, he was continued on steroid taper and commenced on MMF for lupus nephritis. Discussion It can be difficult to differentiate between infective endocarditis and Libman-Sacks endocarditis, especially when there are no existing risk factors. In cases like these antibiotics and steroids can be started at the same time. In our case, we had a young gentleman with no prior medical history who had multiple compounding factors and barriers to fully investigate. Because of his agitation, he required sedation and that allowed the team to perform life saving investigations (renal biopsy and TOE). He ended up having a double metal valve replacement within 8 days post-admission. Multidisciplinary team involvement and close collaboration was of crucial importance in this case to ensure a safe patient outcome. Key learning points • This case illustrates the need to vigorously chase the possibility of concurrent infection in severe new SLE. Close collaboration between multiple specialities to achieve this is key, especially when a patient lacks agency due to their illness.

Culturomics: A promising approach for exploring bacterial diversity in natural fermented milk

Natural fermented milk is an abundant source of microbial resources, some of which possess probiotic properties that can help prevent and treat human diseases. However, many potential probiotics have not yet been isolated using traditional pure-culture methods. Culturomics, a relatively novel approach, can be primarily employed to cultivate and identify previously unknown and uncultured bacteria. Nevertheless, their application in natural fermented milk remains limited. Therefore, the primary objective of this study was to evaluate the bacterial diversity in natural fermented milk and explore its microbial potential. For this purpose, we collected three natural fermented milk samples from Inner Mongolia and subjected them to enrichment culture in blood culture bottles for 14 d. Bacterial isolates were obtained from the samples at 0, 3, 7, and 14 days using two culture temperatures and four media, and full-length 16S rRNA gene sequencing was performed for each sample at each time point. The results revealed that 488 strains were isolated from three samples belonging to 3 phyla, 11 genera, and 27 species. The method using two culture temperatures (20 °C and 30 °C) and two media (MRS and RCM + Vb) yielded most of the isolates after 3, 7, and 14 days of enrichment culture. Furthermore, full-length 16S rRNA gene sequencing confirmed that this method significantly increased bacterial diversity in natural fermented milk. In conclusion, the culturomics method demonstrates great potential for the isolation of a greater number of bacterial species and is advantageous for further exploration of the diversity of culturable microorganisms in natural fermented milk.

Comparative Study of Mesophilic Biomethane Production in Ex Situ Trickling Bed and Bubble Reactors

Biomethane production via biogas upgrading is regarded as a future renewable gas, further boosting the biogas economy. Moreover, when upgrading is realized by the biogas CO2 conversion to CH4 using surplus renewable energy, the process of upgrading becomes a renewable energy storage method. This conversion can be carried out via microorganisms, and has attracted scientific attention, especially under thermophilic conditions. In this study, mesophilic conditions were imposed using a previously developed enriched culture. The enriched culture consisted of the hydrogenotrophic Methanobrevibacter (97% of the Archaea species and 60% of the overall population). Biogas upgrading took place in three lab-scale bioreactors: (a) a 1.2 L bubble reactor (BR), (b) a 2 L trickling bed reactor (TBR) filled with plastic supporting material (TBR-P), and (c) a 1.2 L TBR filled with sintered glass balls (TBR-S). The gas fed into the reactors was a mixture of synthetic biogas and hydrogen, with the H2 to biogas CO2 ratio being 3.7:1, lower than the stoichiometric ratio (4:1). Therefore, the feeding gas mixture did not make it possible for the CH4 content in the biomethane to be more than 97%. The results showed that the BR produced biomethane with a CH4 content of 91.15 ± 1.01% under a gas retention time (GRT) of 12.7 h, while the TBR-P operation resulted in a CH4 content of 90.92 ± 2.15% under a GRT of 6 h. The TBR-S operated at a lower GRT (4 h), yielding an effluent gas richer in CH4 (93.08 ± 0.39%). Lowering the GRT further deteriorated the efficiency but did not influence the metabolic pathway, since no trace of volatile fatty acids was detected. These findings are essential indicators of the process stability under mesophilic conditions.

Isolation and characterization of a resistance Bacillus subtilis for soil stabilization and dust alleviation purposes

Dust poses environmental, geological, health, and economic concerns, and microorganisms can help mitigate these adverse consequences by improving soil properties. Microbial calcium carbonate precipitation (MICP) has been found to be an efficient strategy for increasing soil strength, reducing soil porosity, and preventing erosion; however, severe environmental conditions such as pH and high temperatures may impede this process. To identify the best strain for MICP, 60 bacteria strains were obtained from arid soils using the enrichment culture technique. They were tested for the capacity of calcium carbonate deposition and biocement synthesis in stress environments. Phenotypic characterization indicated that the majority of the bacterial isolates were gram-positive and rod-shaped, with strong catalase and oxidase enzyme activity. Furthermore, MALDI-TOF MS identification revealed that the isolates were from the Bacillus and Pseudomonas genera. Scanning Electron Microscopy (SEM), Energy-dispersive X-ray spectroscopy (EDS), and X-ray diffraction (XRD) were used to analyze the microstructures and composition of bacterial cement. The results represented that B. subtilis isolate S56 has a higher production yield and forms distinctive calcite crystals as a result of fast urease synthesis. B. subtilis isolate S56 can be applied in situ to reduce soil erosion and dust pollution. This study reveals the potential of the B. subtilis S56 strain for soil consolidation and dust prevention in harsh environments and has the prospect of promoting its application in desertification control and ecological restoration.

Improved chickpea growth, physiology, nutrient assimilation and rhizoremediation of hydrocarbons by bacterial consortia.

Soil pollution by petroleum hydrocarbons (PHCs) reduces yield by changing the physico-chemical properties of soil and plants due to PHCs' biotoxicity and persistence. Thus, removing PHCs from the soil is crucial for ecological sustainability. Microbes-assisted phytoremediation is an economical and eco-friendly solution. The current work aimed to develop and use bacterial consortia (BC) for PHCs degradation and plant growth enhancement in hydrocarbon-contaminated soil. Initially, the enriched microbial cultures (that were prepared from PHCs-contaminated soils from five distinct regions) were obtained via screening through microcosm experiments. Afterward, two best microbial cultures were tested for PHCs degradation under various temperature and pH ranges. After culture optimization, isolation and characterization of bacterial strains were done to construct two BC. These constructed BC were tested in a pot experiment for hydrocarbons degradation and chickpea growth in PHCs contaminated soil. Findings revealed that PHCs exerted significant phytotoxic effects on chickpea growth and physiology when cultivated in PHCs contaminated soil, reducing agronomic and physiological traits by 13-29% and 12-43%, respectively. However, in the presence of BC, the phytotoxic impacts of PHCs on chickpea plants were reduced, resulting in up to 24 - 35% improvement in agronomic and physiological characteristics as compared to un-inoculated contaminated controls. Furthermore, the bacterial consortia boosted chickpea's nutritional absorption and antioxidant mechanism. Most importantly, chickpea plants phytoremediated 52% of the initial PHCs concentration; however, adding BC1 and BC2 with chickpea plants further increased this removal and remediated 74% and 80% of the initial PHCs concentration, respectively. In general, BC2 outperformed BC1 (with few exceptions) in promoting plant growth and PHCs elimination. Therefore, using multi-trait BC for PHCs degradation and plant growth improvement under PHCs stress may be an efficient and environmentally friendly strategy to deal with PHCs pollution and toxicity.

Study of biotechnological potential of hydrocarbon-oxidizing bacteria from oil-contaminated soils of the Uzen oil field

Background: Oil-contaminated soils are one of the significant environmental concerns in Western Kazakhstan. Cleaning soils from oil contamination is becoming extremely important. Biological soil treatment, which uses hydrocarbon-oxiding bacteria for microbiological remediation, is a more environmentally friendly and delicate method of soil treatment than any other currently used technology (physical, chemical, biological). One of the important aspects of microbiological remediation is the use of bacteria endomimic to the treated soil. This guarantees the most effective soil purification, as the bacteria perform in relatively optimal conditions for themselves. The soil of the Mangistau region, especially the coastal regions, is distinctive due to its high mineralisation and low moisture content, which have helped to form a specific microflora that is adapted to these conditions. Aim: The study aims to search and isolate highly effective hydrocarbon-oxidizing bacteria that are native to highly mineralized soils of the Uzen oil field. Materials and methods: The research employs a variety of analytical techniques, including water chemistry analysis and infrared spectrometry, as well as the microbiological culture of aerobic bacteria on liquid and dense media in environments (temperature and salinity) that closely resemble their native ecotopes. Results: Four enrichment cultures of hydrocarbon-oxidizing bacteria were obtained, and three pure cultures of oil-oxidizing bacteria were isolated. Their hydrocarbon-oxidizing efficiency has been studied. On the basis of heavy, extremely viscous paraffinic oil from the Uzen field, their hydrocarbon-oxidizing efficiency was examined. Conclusion: In this study, active enrichment cultures of halophilic hydrocarbon-oxidizing bacteria as well as active accumulative cultures of halophilic and moderately thermophilic aerobic bacteria were obtained. These bacteria are capable of oxidising a wide range of hydrocarbons, including high-molecular polycyclic and sulfur-containing compounds. Their high biotechnological potential will be studied in further studies.

Ukrainian Musical Culture: Integrative Component

The purpose of the research to study the current state of Ukrainian musical culture and to prove the importance of its European integration. Research methodology. To achieve the set goals, the following research methods were used: historical - to study the emergence, formation and development of Ukrainian musical culture; systematic method – for a comprehensive study of the musical culture of Ukraine; cultural – which reveals the complexity of socio-cultural processes and phenomena; axiological method – which allowed to investigate the phenomena of mass culture in view of their valuable nature. Scientific novelty the article consists in the theoretical and methodological substantiation of the influence of the European space on the Europeanisation of Ukrainian culture in Ukraine and beyond. Conclusions. The disclosure of the concept of Ukrainian culture showed that the constituent parts of any culture are the people with their customs, beliefs, traditions, as well as innovations in this field, which are closely intertwined with the past heritage. Enrichment of culture is possible only if the experience of past generations is successfully combined with modern manifestations of cultural development. Having considered the main factors and determining factors of national culture, we proved that art itself, in particular music, are determining factors of culture. It is music that is capable of enriching and elevating the spiritual value of the Ukrainian people, because the feeling of beauty aesthetically directs the personality, contributes to value orientations and conscious being. The interpenetration of different cultures and generations leads to the development of innovations, while the use of some of the latest technologies loses the meaning of national culture. It is necessary to enrich the spiritual culture of Ukraine – as an important factor in the development of a modern, educated personality. National musical heritage is the primary source of formation of national consciousness and identity. Integration processes and cultural dialogue are a necessary condition for the development of Ukrainian musical culture, as well as any culture in general. A dialogue that involves active interaction, mutual enrichment of equal cultural subjects, a desire to understand one's own and others' peculiarities and uniqueness.

The acceptability of implementation of group B Streptococcus testing: Perspectives from women and health professionals in the GBS3 trial: A qualitative study

ObjectiveTo determine the acceptability of different methods of routine testing for group B Streptococcus (GBS) colonisation to pregnant women and health care professionals (HCPs), and to examine barriers and facilitators to their implementation. DesignQualitative study, embedded in a cluster randomised trial SettingFour NHS maternity units participating in the GBS3 Trial: two conducting routine antenatal enriched culture medium (ECM) testing; and two using routine rapid intrapartum testing.Sample39 women and 25 HCPs purposively sampled to ensure representation of women with various birthing experiences and different professions.MethodsWomen were interviewed approximately 12 weeks postpartum by telephone or online video call, using a semi-structured topic guide. HCPs were interviewed during the testing period of the trial. Interviews were transcribed for thematic analysis and summarised using the framework method. ResultsFour categories of interest emerged: (1) views of routine testing; (2) acceptability of the testing procedure; (3) preferences on the types of test; (4) improving the testing procedure. Routine GBS testing was well received by both women and HCPs. Most participants found the procedure acceptable and were willing to receive the offer of testing in the future. Preferences for different testing methods varied, with participants emphasising the importance of evidence and informed choice. ConclusionsRoutine GBS testing is acceptable to most women and HCPs. Areas for consideration and the practicalities of implementing testing in maternity services are highlighted.

Mycobacterium tuberculosis complex sample processing by mechanical lysis, an essential step for reliable whole genome sequencing

Mycobacterium tuberculosis complex (MTBC) whole genome sequencing (WGS) turnaround time and WGS success rates are highly influenced by DNA extraction protocols even from cultures. Efficient mycobacterial lysis is crucial for obtaining sufficient DNA from cultures to facilitate reliable genomic drug susceptibility prediction and accurate genotyping with WGS. We compared four DNA extraction protocols from BD BACTEC™ Mycobacterial Growth Indicator Tubes (MGIT) for WGS with a focus on the lysis step: protocol A) column-based protocol without mechanical lysis; protocol B) an adapted protocol including a bead beating step; protocol C) DNA extraction from primary received cultures using bead beating: and protocol D) DNA extraction from pre MGIT-positive (enriched) cultures. Protocol B increased DNA yield approximately 60-fold, and significantly improved the sequencing success rate. The increased yield also allowed DNA extraction from primary cultures with high success rates (protocol C). Additionally, by using pre-positive enriched MGIT cultures, we demonstrated that bead beating opens the possibility of reliable WGS up to five days before a MGIT tube would be flagged positive (protocol D). The most optimal bead beating-based DNA extraction was also evaluated for Nanopore sequencing. Shortening bead beating duration to 15 s resulted in longer read lengths (N50 from 1.4 kb to 2.6 kb) while still providing efficient lysis. Furthermore, AmpureXP bead beating-based DNA capture / purification proved to be as efficient as Qiagen column-based DNA extraction, further simplifying and shortening the DNA extraction protocol. Adding a mechanical lysis step to our routine MTBC DNA extraction protocol has allowed us to reduce the turnaround time while maintaining DNA quality sequencing success rates.

Deciphering Hyperammonia-Producing Bacteria (HAB) in the Rumen of Water Buffaloes (Bubalus bubalis) and Their Inhibition through Plant Extracts and Essential Oils

Hyperammonia-producing bacteria (HAB) are a class of microbes present in the stomach of ruminants, responsible for the rapid rate of ammonia production from protein degradation beyond the capacity of these animals for their utilization. Thus, ruminant nutritionists are interested in decreasing ruminal protein degradation and ammonia genesis by focusing on controlling the activity of HAB. The investigations of the present study were carried out to determine predominant hyperammonia-producing bacteria in the rumen of buffaloes, their isolation and characterization, as well as the inhibition of these isolates with various sources of plant secondary compounds (tannins, saponins, and essential oils). Studies employing high-throughput sequencing of amplicons of the 16S rRNA gene from genomic DNA recovered from enrichment culture of HAB of buffalo rumina indicated that, at the phylum level, Proteobacteria (61.1 to 68.2%) was the most predominant HAB. Acidaminococcus was most predominant among the identified genera. In vitro experiments were conducted with enrichment culture of buffalo rumen contents incubated with different types of feed additives such as essential oils (eucalyptus oil, lemon grass oil, and clove oil) and extracts of plants (Sapindus mukorossi fruits and Ficus bengalensis leaves), each at graded dose levels. The reduction in ammonia production by clove and lemon grass oils was evident due to the presence of major bioactive compounds, especially eugenol and limonene, which have strong antimicrobial activity. However, clove oil and Indian soapberry/reetha (Sapindus mukorossi) fruit were found to be promising and effective in reducing the growth, protease production, and ammonia production of HAB culture.

A dataset for developing proteomic tools for pathogen detection via differential cell lysis of whole blood samples

This data descriptor presents a curated dataset for pathogen detection and identification (Staphylococcus aureus, Pseudomonas aeruginosa, and Candida albicans) directly from whole-blood samples. The dataset was created using differential cell lysis combined with rapid extraction, digestion, and mass spectrometry-based proteomics. Our method offers a rapid diagnostic alternative to traditional culture, enabling timely disease management, such as sepsis. Highlighting our dataset’s uniqueness, it features a three-tier structure: Spectral Libraries of Pathogens for identifying peptide peaks for putative biomarkers; Spiked pathogen in blood MS data for biomarker panel optimization through varied concentration samples; and Parallel Reaction Monitoring (PRM) data from sepsis patients for validating our biomarker panel, achieving 83.3% sensitivity within seven hours without microbial enrichment culture. This dataset serves as a comprehensive reference for bioinformatic tool development and biomarker panel proposals, advancing microbial detection, antimicrobial resistance, and epidemiological studies.

Environmental monitoring of Listeria monocytogenes contamination in dairy processing facilities combining culturing technique and molecular methods

Objectiveof the present study was to investigate Listeria contamination in Sardinian sheep cheese-making plants using culture and DNA-based methods. Food and environmental samples were collected from different surfaces of 14 facilities. Samples were collected using pre-moistened sponge swabs. Each site was sampled using two sponges used one for microbiological detection and the other for extraction of total DNA. The DNA was submitted to real time PCR targeting the prs gene for Listeria spp., the gene inlA for Listeria monocytogenes and the gene iap for Listeria innocua. DNA metabarcoding was also conducted on the total DNA. Overall were collected 254 environmental and 36 food samples. The prevalence of Listeria spp. and Listeria monocytogenes was 18.6% and 10.0%, respectively. Samples enrichment coupled with quantitative PCR (qPCR) showed a greater sensitivity than the conventional culture method with an overall prevalence of 12.1% for L. monocytogenes. Both, qPCR and ribosomal DNA-metabarcoding conducted on total DNA extracted from the sponges performed poorly in comparison to the culture-based method detecting the presence of Listeria genus in as little as 7 (2.4%) and 4 (1.4%) samples respectively. The present study supports the use of a culture enrichment coupled with qPCR for routine monitoring of contamination in food premises.