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25 pages, 6067 KiB

Open AccessArticle

Essential Oils as Alternative Biocides for the Preservation of Waterlogged Archaeological Wood

by Federica Antonelli, Marco Bartolini, Marie-Laure Plissonnier, Alfonso Esposito, Giulia Galotta, Sandra Ricci, Barbara Davidde Petriaggi, Cristian Pedone, Antonella Di Giovanni, Silvano Piazza, Francesca Guerrieri and Manuela Romagnoli

Microorganisms 2020, 8(12), 2015; https://doi.org/10.3390/microorganisms8122015 - 16 Dec 2020

Cited by 25 | Viewed by 3310

Abstract

Waterlogged archaeological wood is exposed to a high risk of biological degradation during the post-excavation phases of storage and restoration. For this reason, often biocides must be used to preserve wooden remains. In the present work three essential oils (cinnamon, wild thyme, and [...] Read more.

Waterlogged archaeological wood is exposed to a high risk of biological degradation during the post-excavation phases of storage and restoration. For this reason, often biocides must be used to preserve wooden remains. In the present work three essential oils (cinnamon, wild thyme, and common thyme) were tested as possible alternative biocides to use in the preservation of waterlogged archaeological wood. The oils were first tested in vitro to establish the minimum inhibitory concentration (MIC) and to evaluate the biocidal activity on selected fungal strains. Then, the established MIC was applied on waterlogged archaeological wood samples and during an actual restoration treatment. The effectiveness of the oils was evaluated through cultural analyses, ATP quantification, and next-generation sequencing. The results showed that the oils caused a significant decrease in the vitality of fungal mycelia grown in vitro and of the microbiota present in treated wood and storage water. Furthermore, an influence on the composition of the bacterial communities of treated wood samples was observed. Although further tests are needed to evaluate interferences with the materials used during restoration procedures, essential oils could be considered as a possible alternative to the currently used biocide. Full article

(This article belongs to the Special Issue New Methods in Microbial Research)

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21 pages, 2337 KiB

Open AccessArticle

Tn6603, a Carrier of Tn5053 Family Transposons, Occurs in the Chromosome and in a Genomic Island of Pseudomonas aeruginosa Clinical Strains

by Vaheesan Rajabal, Vilma A. Stanisich and Steve Petrovski

Microorganisms 2020, 8(12), 1997; https://doi.org/10.3390/microorganisms8121997 - 15 Dec 2020

Cited by 1 | Viewed by 2331

Abstract

Transposons of the Pseudomonasaeruginosa accessory gene pool contribute to phenotype and to genome plasticity. We studied local P. aeruginosa strains to ascertain the encroachment of mer-type res site hunter transposons into clinical settings and their associations with other functional modules. Five [...] Read more.

Transposons of the Pseudomonasaeruginosa accessory gene pool contribute to phenotype and to genome plasticity. We studied local P. aeruginosa strains to ascertain the encroachment of mer-type res site hunter transposons into clinical settings and their associations with other functional modules. Five different Tn5053 family transposons were detected, all chromosomal. Some were solitary elements; one was in res of Tn1013^#, a relative of a reported carrier of int-type res site hunters (class 1 integrons), but most were in res of Tn6603, a new Tn501-related transposon of unknown phenotype. Most of the Tn6603::Tn elements, and some Tn6603 and Tn6603::Tn elements found in GenBank sequences, were at identical sites in an hypothetical gene of P. aeruginosa genomic island PAGI-5v. The island in clonally differing strains was at either of two tRNA^Lys loci, suggesting lateral transfer to these sites. This observation is consistent with the membership of the prototype PAGI-5 island to the ICE family of mobile genetic elements. Additionally, the res site hunters in the nested transposons occupied different positions in the Tn6603 carrier. This suggested independent insertion events on five occasions at least. Tn5053 family members that were mer-/tni-defective were found in Tn6603- and Tn501-like carriers in GenBank sequences of non-clinical Pseudomonas spp. The transposition events in these cases presumably utilized tni functions in trans, as can occur with class 1 integrons. We suggest that in the clinical context, P. aeruginosa strains that carry Tn6603 alone or in PAGI-5v can serve to disseminate functional res site hunters; these in turn can provide the requisite trans-acting tni functions to assist in the dissemination of class 1 integrons, and hence of their associated antibiotic resistance determinants. Full article

(This article belongs to the Special Issue New Methods in Microbial Research)

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22 pages, 4216 KiB

Open AccessArticle

Evaluating the Effects of Disinfectants on Bacterial Biofilms Using a Microfluidics Flow Cell and Time-Lapse Fluorescence Microscopy

by Milos Legner, James Jonkman and Dean Swift

Microorganisms 2020, 8(11), 1837; https://doi.org/10.3390/microorganisms8111837 - 22 Nov 2020

Cited by 1 | Viewed by 3259

Abstract

A commercially available microfluidics flow cell was utilized together with widefield fluorescence microscopy to evaluate the effects of disinfectants on bacterial strains. The flow cell’s inner surface supports the formation of biofilms of numerous bacterial species. The modular setup of the flow cell [...] Read more.

A commercially available microfluidics flow cell was utilized together with widefield fluorescence microscopy to evaluate the effects of disinfectants on bacterial strains. The flow cell’s inner surface supports the formation of biofilms of numerous bacterial species. The modular setup of the flow cell accessories allows connection to syringes, pumps and collection vials, facilitating aseptic experiments in a controlled fluidics environment which can be documented with precisely timed microscopy imaging. The flow cell is inoculated with a suspension of bacteria in a nutrient medium and incubated for several days allowing bacterial cells to form a biofilm. Shortly before performing an assay, the biofilm is labelled with a dual-fluorescent DNA probe which distinguishes unharmed and damaged bacteria. Then a disinfectant sample (or control) is gently injected and time-lapse imaging is used for quantifying the course of bacterial biomass response. We use a simplified widefield microscopy method that allows intensive recording and quantification of time series of two-dimensional frames for tracking the course of disinfectant action on a variety of microbial strains. This procedure has potential for the rapid evaluation of novel products. Full article

(This article belongs to the Special Issue New Methods in Microbial Research)

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18 pages, 383 KiB

Open AccessArticle

Clustering on Human Microbiome Sequencing Data: A Distance-Based Unsupervised Learning Model

by Dongyang Yang and Wei Xu

Microorganisms 2020, 8(10), 1612; https://doi.org/10.3390/microorganisms8101612 - 20 Oct 2020

Cited by 6 | Viewed by 3895

Abstract

Modeling and analyzing human microbiome allows the assessment of the microbial community and its impacts on human health. Microbiome composition can be quantified using 16S rRNA technology into sequencing data, which are usually skewed and heavy-tailed with excess zeros. Clustering methods are useful [...] Read more.

Modeling and analyzing human microbiome allows the assessment of the microbial community and its impacts on human health. Microbiome composition can be quantified using 16S rRNA technology into sequencing data, which are usually skewed and heavy-tailed with excess zeros. Clustering methods are useful in personalized medicine by identifying subgroups for patients stratification. However, there is currently a lack of standardized clustering method for the complex microbiome sequencing data. We propose a clustering algorithm with a specific beta diversity measure that can address the presence-absence bias encountered for sparse count data and effectively measure the sample distances for sample stratification. Our distance measure used for clustering is derived from a parametric based mixture model producing sample-specific distributions conditional on the observed operational taxonomic unit (OTU) counts and estimated mixture weights. The method can provide accurate estimates of the true zero proportions and thus construct a precise beta diversity measure. Extensive simulation studies have been conducted and suggest that the proposed method achieves substantial clustering improvement compared with some widely used distance measures when a large proportion of zeros is presented. The proposed algorithm was implemented to a human gut microbiome study on Parkinson’s diseases to identify distinct microbiome states with biological interpretations. Full article

(This article belongs to the Special Issue New Methods in Microbial Research)

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18 pages, 6333 KiB

Open AccessArticle

RBUD: A New Functional Potential Analysis Approach for Whole Microbial Genome Shotgun Sequencing

by Zhikai Xing, Yunting Zhang, Meng Li, Chongye Guo and Shuangli Mi

Microorganisms 2020, 8(10), 1563; https://doi.org/10.3390/microorganisms8101563 - 10 Oct 2020

Cited by 1 | Viewed by 2585

Abstract

Whole metagenome shotgun sequencing is a powerful approach to detect the functional potential of microbial communities. Currently, the read-based metagenomics profiling for established database (RBED) method is one of the two kinds of conventional methods for species and functional annotations. However, the databases, [...] Read more.

Whole metagenome shotgun sequencing is a powerful approach to detect the functional potential of microbial communities. Currently, the read-based metagenomics profiling for established database (RBED) method is one of the two kinds of conventional methods for species and functional annotations. However, the databases, which are established based on test samples or specific reference genomes or protein sequences, limit the coverage of global microbial diversity. The other assembly-based metagenomics profiling for unestablished database (ABUD) method has a low utilization rate of reads, resulting in a lot of biological information loss. In this study, we proposed a new method, read-based metagenomics profiling for unestablished database (RBUD), based on Metagenome Database of Global Microorganisms (MDGM), to solve the above problems. To evaluate the accuracy and effectiveness of our method, the intestinal bacterial composition and function analyses were performed in both avian colibacillosis chicken cases and type 2 diabetes mellitus patients. Comparing to the existing methods, RBUD is superior in detecting proteins, percentage of reads mapping and ontological similarity of intestinal microbes. The results of RBUD are in better agreement with the classical functional studies on these two diseases. RBUD also has the advantages of fast analysis speed and is not limited by the sample size. Full article

(This article belongs to the Special Issue New Methods in Microbial Research)

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12 pages, 3036 KiB

Open AccessArticle

Genotypic and Phenotypic Characterization of Treponema phagedenis from Bovine Digital Dermatitis

by Hector M. Espiritu, Lovelia L. Mamuad, Su-jeong Jin, Seon-ho Kim, Seok-won Kwon, Sang-suk Lee, Sang-myeong Lee and Yong-il Cho

Microorganisms 2020, 8(10), 1520; https://doi.org/10.3390/microorganisms8101520 - 02 Oct 2020

Cited by 5 | Viewed by 2266

Abstract

This study aimed to isolate and characterize Treponema spp. from bovine digital dermatitis (BDD)-infected dairy cattle. Seven isolates were characterized in this study. Isolates exhibited slow growth, and colonies penetrated the agar and exhibited weak β-hemolysis. Round bodies were observed in old and [...] Read more.

This study aimed to isolate and characterize Treponema spp. from bovine digital dermatitis (BDD)-infected dairy cattle. Seven isolates were characterized in this study. Isolates exhibited slow growth, and colonies penetrated the agar and exhibited weak β-hemolysis. Round bodies were observed in old and antibiotic-treated cultures. Cells ranged from 9–12 µm in length, 0.2–2.5 µm in width, and were moderately spiraled. The 16S rRNA analysis revealed the isolates as Treponema phagedenis with >99% sequence homology. Isolates had alkaline phosphatase, acid phosphatase, β-galactosidase, N-acetyl-β-glucosaminidase, esterase (C4), esterase lipase (C8), naphthol-AS-BI-phosphohydrolase, and β-glucuronidase activities. Low concentrations of ampicillin, erythromycin, and tetracycline were required to inhibit the growth of isolates. Formic, acetic, and butyric acids were produced, while propionic acid was significantly utilized, indicating its essentiality for treponemal growth. The isolates shared the same characteristics and, therefore, were considered as a single strain. Isolate HNL4 was deposited as a representative isolate (Treponema phagedenis KS1). The average nucleotide identity of strain KS1 showed a small difference with the human strain (99.14%) compared with bovine strain (99.72%). This study was the first to isolate and characterize Treponema phagedenis from BDD in Korea and, hence, it delivered pathogenicity-related insights and provided valuable information that can be used for the management of BDD. Full article

(This article belongs to the Special Issue New Methods in Microbial Research)

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14 pages, 3242 KiB

Open AccessArticle

Phylogenetic Assignment of the Fungicolous Hypoxylon invadens (Ascomycota, Xylariales) and Investigation of its Secondary Metabolites

by Kevin Becker, Christopher Lambert, Jörg Wieschhaus and Marc Stadler

Microorganisms 2020, 8(9), 1397; https://doi.org/10.3390/microorganisms8091397 - 11 Sep 2020

Cited by 11 | Viewed by 2890

Abstract

The ascomycete Hypoxylon invadens was described in 2014 as a fungicolous species growing on a member of its own genus, H.fragiforme, which is considered a rare lifestyle in the Hypoxylaceae. This renders H.invadens an interesting target in our efforts to [...] Read more.

The ascomycete Hypoxylon invadens was described in 2014 as a fungicolous species growing on a member of its own genus, H.fragiforme, which is considered a rare lifestyle in the Hypoxylaceae. This renders H.invadens an interesting target in our efforts to find new bioactive secondary metabolites from members of the Xylariales. So far, only volatile organic compounds have been reported from H.invadens, but no investigation of non-volatile compounds had been conducted. Furthermore, a phylogenetic assignment following recent trends in fungal taxonomy via a multiple sequence alignment seemed practical. A culture of H.invadens was thus subjected to submerged cultivation to investigate the produced secondary metabolites, followed by isolation via preparative chromatography and subsequent structure elucidation by means of nuclear magnetic resonance (NMR) spectroscopy and high-resolution mass spectrometry (HR-MS). This approach led to the identification of the known flaviolin (1) and 3,3-biflaviolin (2) as the main components, which had never been reported from the order Xylariales before. Assessment of their antimicrobial and cytotoxic effects via a panel of commonly used microorganisms and cell lines in our laboratory did not yield any effects of relevance. Concurrently, genomic DNA from the fungus was used to construct a multigene phylogeny using ribosomal sequence information from the internal transcribed spacer region (ITS), the 28S large subunit of ribosomal DNA (LSU), and proteinogenic nucleotide sequences from the second largest subunit of the DNA-directed RNA polymerase II (RPB2) and β-tubulin (TUB2) genes. A placement in a newly formed clade with H.trugodes was strongly supported in a maximum-likelihood (ML) phylogeny using sequences derived from well characterized strains, but the exact position of said clade remains unclear. Both, the chemical and the phylogenetic results suggest further inquiries into the lifestyle of this unique fungus to get a better understanding of both, its ecological role and function of its produced secondary metabolites hitherto unique to the Xylariales. Full article

(This article belongs to the Special Issue New Methods in Microbial Research)

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7 pages, 796 KiB

Open AccessCommunication

Key Bacteria in the Gut Microbiota Network for the Transition between Sedentary and Active Lifestyle

by Nazareth Castellanos, Gustavo G. Diez, Carmen Antúnez-Almagro, Carlo Bressa, María Bailén, Rocío González-Soltero, Margarita Pérez and Mar Larrosa

Microorganisms 2020, 8(5), 785; https://doi.org/10.3390/microorganisms8050785 - 24 May 2020

Cited by 10 | Viewed by 4298

Abstract

Physical activity modifies the gut microbiota, exerting health benefits on the host; however, the specific bacteria associated with exercise are not yet known. In this work, we propose a novel method, based on hierarchical topology, to study the differences between the microbiota of [...] Read more.

Physical activity modifies the gut microbiota, exerting health benefits on the host; however, the specific bacteria associated with exercise are not yet known. In this work, we propose a novel method, based on hierarchical topology, to study the differences between the microbiota of active and sedentary lifestyles, and to identify relevant bacterial taxa. Our results show that the microbiota network found in active people has a significantly higher overall efficiency and higher transmissibility rate. We also identified key bacteria in active and sedentary networks that could be involved in the conversion of an active microbial network to a sedentary microbial network and vice versa. Full article

(This article belongs to the Special Issue New Methods in Microbial Research)

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10 pages, 3654 KiB

Open AccessCommunication

Quantifying Plant-Borne Carbon Assimilation by Root-Associating Bacteria

by Spenser Waller, Stacy L. Wilder, Michael J. Schueller, Alexandra B. Housh and Richard A. Ferrieri

Microorganisms 2020, 8(5), 700; https://doi.org/10.3390/microorganisms8050700 - 10 May 2020

Cited by 6 | Viewed by 3314

Abstract

Herbaspirillum seropedicae is a rhizobacteria that occupies a specialized ecological niche in agriculture. As an endophyte and prolific grass root colonizer it has the potential to promote plant growth, enhancing crop yield in many cereal crops. While the mechanisms for plant growth promotion [...] Read more.

Herbaspirillum seropedicae is a rhizobacteria that occupies a specialized ecological niche in agriculture. As an endophyte and prolific grass root colonizer it has the potential to promote plant growth, enhancing crop yield in many cereal crops. While the mechanisms for plant growth promotion are controversial, the one irrefutable fact is these microorganisms rely heavily on plant-borne carbon as their main energy source in support of their biological functions. Unfortunately, the tools and technology enabling researchers to trace carbon exchange between plants and the microorganisms associating with them has been limiting. Here, we demonstrate that radioactive ¹¹CO₂ administered to intact maize leaves with translocation of ¹¹C-photosynthates to roots can provide a ‘traceable’ source of carbon whose assimilation by microbial organisms can be quantified with enormous sensitivity. Fluorescence root imaging of RAM10, a green fluorescent protein (GFP) reporting strain of H. seropedicae, was used to identify regions of high microbial colonization. Microbes were mechanically removed from these regions via sonication in saline solution and extracts were subjected to fluorescence measurement and gamma counting to correlate carbon-11 atoms with numbers of colony forming units. The method has potential to translate to other microorganisms provided they possess an optical reporting trait. Full article

(This article belongs to the Special Issue New Methods in Microbial Research)

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23 pages, 4393 KiB

Open AccessArticle

Framework as a Service, FaaS: Personalized Prebiotic Development for Infants with the Elements of Time and Parametric Modelling of In Vitro Fermentation

by Ka-Lung Lam, Wai-Yin Cheng, Fan Yang, Shaoling Lin, Lijun You, Jiachi Chiou, Hoi-Shan Kwan and Peter Chi-Keung Cheung

Microorganisms 2020, 8(5), 623; https://doi.org/10.3390/microorganisms8050623 - 25 Apr 2020

Viewed by 2537

Abstract

We proposed a framework with parametric modeling to obtain biological relevant parameters from the total probiotic growth pattern and metabolite production curves. The lag phase, maximum increase rate, and maximum capacity were obtained via a 205-h exploratory In vitro fermentation of a library [...] Read more.

We proposed a framework with parametric modeling to obtain biological relevant parameters from the total probiotic growth pattern and metabolite production curves. The lag phase, maximum increase rate, and maximum capacity were obtained via a 205-h exploratory In vitro fermentation of a library of 13 structural-characterized prebiotic candidates against an exclusively breastfed infant fecal inoculum. We also conducted 16S rRNA amplicon sequencing of the infant fecal inoculum. Moreover, we introduce a robust composite metabolite-based indicator that reflects the eubiosis/dysbiosis of microbiota to complement the conventional microbial markers. In terms of short-chain fatty acid, we discovered that polymeric beta-glucans from barley demonstrated potential as prebiotic candidates, while alpha-glucans as glycogen showed the least dissolved ammonia production. In terms of total probiotic, beta-glucans from oat and mushroom sclerotia of Pleurotus tuber-regium showed comparable sustainability when compared to alpha-glucans after 48 h. Being classical prebiotic, galacto-oligosaccharides gave the second-highest metabolite-based indicator, followed by lactose. While limited improvement could be made to lactose and oligosaccharides, polymeric beta-glucans from barley avails more capacity for novel prebiotic development, such as structural modification. We anticipate that more similar parallel screening with the element of time and parametric modeling will provide more novel insights. Full article

(This article belongs to the Special Issue New Methods in Microbial Research)

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Review

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28 pages, 398 KiB

Open AccessReview

Current Pretreatment/Cell Disruption and Extraction Methods Used to Improve Intracellular Lipid Recovery from Oleaginous Yeasts

by Muhammad Fakhri Zainuddin, Chong Kar Fai, Arbakariya B. Ariff, Leonardo Rios-Solis and Murni Halim

Microorganisms 2021, 9(2), 251; https://doi.org/10.3390/microorganisms9020251 - 27 Jan 2021

Cited by 38 | Viewed by 6480

Abstract

The production of lipids from oleaginous yeasts involves several stages starting from cultivation and lipid accumulation, biomass harvesting and finally lipids extraction. However, the complex and relatively resistant cell wall of yeasts limits the full recovery of intracellular lipids and usually solvent extraction [...] Read more.

The production of lipids from oleaginous yeasts involves several stages starting from cultivation and lipid accumulation, biomass harvesting and finally lipids extraction. However, the complex and relatively resistant cell wall of yeasts limits the full recovery of intracellular lipids and usually solvent extraction is not sufficient to effectively extract the lipid bodies. A pretreatment or cell disruption method is hence a prerequisite prior to solvent extraction. In general, there are no recovery methods that are equally efficient for different species of oleaginous yeasts. Each method adopts different mechanisms to disrupt cells and extract the lipids, thus a systematic evaluation is essential before choosing a particular method. In this review, mechanical (bead mill, ultrasonication, homogenization and microwave) and nonmechanical (enzyme, acid, base digestions and osmotic shock) methods that are currently used for the disruption or permeabilization of oleaginous yeasts are discussed based on their principle, application and feasibility, including their effects on the lipid yield. The attempts of using conventional and “green” solvents to selectively extract lipids are compared. Other emerging methods such as automated pressurized liquid extraction, supercritical fluid extraction and simultaneous in situ lipid recovery using capturing agents are also reviewed to facilitate the choice of more effective lipid recovery methods. Full article

(This article belongs to the Special Issue New Methods in Microbial Research)

12 pages, 231 KiB

Open AccessReview

The Role of Urban Wastewater in the Environmental Transmission of Antimicrobial Resistance: The Current Situation in Italy (2010–2019)

by Francesco Triggiano, Carla Calia, Giusy Diella, Maria Teresa Montagna, Osvalda De Giglio and Giuseppina Caggiano

Microorganisms 2020, 8(10), 1567; https://doi.org/10.3390/microorganisms8101567 - 12 Oct 2020

Cited by 22 | Viewed by 2447

Abstract

Scientific studies show that urban wastewater treatment plants (UWWTP) are among the main sources of release of antibiotics, antibiotic resistance genes (ARG) and antibiotic-resistant bacteria (ARB) into the environment, representing a risk to human health. This review summarizes selected publications from 1 January [...] Read more.

Scientific studies show that urban wastewater treatment plants (UWWTP) are among the main sources of release of antibiotics, antibiotic resistance genes (ARG) and antibiotic-resistant bacteria (ARB) into the environment, representing a risk to human health. This review summarizes selected publications from 1 January 2010 to 31 December 2019, with particular attention to the presence and treatment of ARG and ARB in UWWTPs in Italy. Following a brief introduction, the review is divided into three sections: (i) phenotypic assessment (ARB) and (ii) genotypic assessment (ARG) of resistant microorganisms, and (iii) wastewater treatment processes. Each article was read entirely to extract the year of publication, the geographical area of the UWWTP, the ARB and ARG found, and the type of disinfection treatment used. Among the ARB, we focused on the antibiotic resistance of Escherichia coli, Klebsiella pneumoniae, and Enterococci in UWWTP. The results show that the information presented in the literature to date is not exhaustive; therefore, future scientific studies at the national level are needed to better understand the spread of ARB and ARG, and also to develop new treatment methods to reduce this spread. Full article

(This article belongs to the Special Issue New Methods in Microbial Research)

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