Influence of Substrate Concentration on the Culturability of Heterotrophic Soil Microbes Isolated by High-Throughput Dilution-to-Extinction Cultivation

The vast majority of microbes inhabiting oligotrophic shallow subsurface soil environments have not been isolated or studied under controlled laboratory conditions. In part, the challenges associated with isolating shallow subsurface microbes may persist because microbes in deeper soils are adapted to low nutrient availability or quality. Here, we use high-throughput dilution-to-extinction culturing to isolate shallow subsurface microbes from a conifer forest in Arizona, USA. We hypothesized that the concentration of heterotrophic substrates in microbiological growth medium would affect which microbial taxa were culturable from these soils. To test this, we diluted cells extracted from soil into one of two custom-designed defined growth media that differed by 100-fold in the concentration of amino acids and organic carbon. Across the two media, we isolated a total of 133 pure cultures, all of which were classified as Actinobacteria or Alphaproteobacteria The substrate availability dictated which actinobacterial phylotypes were culturable but had no significant effect on the culturability of Alphaproteobacteria We isolated cultures that were representative of the most abundant phylotype in the soil microbial community (Bradyrhizobium spp.) and representatives of five of the top 10 most abundant Actinobacteria phylotypes, including Nocardioides spp., Mycobacterium spp., and several other phylogenetically divergent lineages. Flow cytometry of nucleic acid-stained cells showed that cultures isolated on low-substrate medium had significantly lower nucleic acid fluorescence than those isolated on high-substrate medium. These results show that dilution-to-extinction is an effective method to isolate abundant soil microbes and that the concentration of substrates in culture medium influences the culturability of specific microbial lineages.IMPORTANCE Isolating environmental microbes and studying their physiology under controlled conditions are essential aspects of understanding their ecology. Subsurface ecosystems are typically nutrient-poor environments that harbor diverse microbial communities-the majority of which are thus far uncultured. In this study, we use modified high-throughput cultivation methods to isolate subsurface soil microbes. We show that a component of whether a microbe is culturable from subsurface soils is the concentration of growth substrates in the culture medium. Our results offer new insight into technical approaches and growth medium design that can be used to access the uncultured diversity of soil microbes.Open access journalThis item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

Characterization of Diverse Anelloviruses, Cressdnaviruses, and Bacteriophages in the Human Oral DNA Virome from North Carolina (USA)

The diversity of viruses identified from the various niches of the human oral cavity—from saliva to dental plaques to the surface of the tongue—has accelerated in the age of metagenomics. This rapid expansion demonstrates that our understanding of oral viral diversity is incomplete, with only a few studies utilizing passive drool collection in conjunction with metagenomic sequencing methods. For this pilot study, we obtained 14 samples from healthy staff members working at the Duke Lemur Center (Durham, NC, USA) to determine the viral diversity that can be identified in passive drool samples from humans. The complete genomes of 3 anelloviruses, 9 cressdnaviruses, 4 Caudoviricetes large bacteriophages, 29 microviruses, and 19 inoviruses were identified in this study using high-throughput sequencing and viral metagenomic workflows. The results presented here expand our understanding of the vertebrate-infecting and microbe-infecting viral diversity of the human oral virome in North Carolina (USA)

Identification of a novel polyomavirus in wild Sonoran Desert rodents of the family Heteromyidae

Rodents are the largest and most diverse group of mammals. Covering a wide range of structural and functional adaptations, rodents successfully occupy virtually every terrestrial habitat, and they are often found in close association with humans, domestic animals, and wildlife. Although a significant amount of research has focused on rodents’ prominence as known reservoirs of zoonotic viruses, there has been less emphasis on the viral ecology of rodents in general. Here, we utilized a viral metagenomics approach to investigate polyomaviruses in wild rodents from the Baja California peninsula, Mexico, using fecal samples. We identified a novel polyomavirus in fecal samples from two rodent species, a spiny pocket mouse (Chaetodipus spinatus) and a Dulzura kangaroo rat (Dipodomys simulans). These two polyomaviruses represent a new species in the genus Betapolyomavirus. Sequences of this polyomavirus cluster phylogenetically with those of other rodent polyomaviruses and two other non-rodent polyomaviruses (WU and KI) that have been identified in the human respiratory tract. Through our continued work on seven species of rodents, we endeavor to explore the viral diversity associated with wild rodents on the Baja California peninsula and expand on current knowledge of rodent viral ecology and evolution.12 month embargo; first published: 15 September 2023This item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

Short communication: Macrofungal diversity in Mt. Makiling forest reserve, Laguna, Philippines: With floristic update on roadside samples in Makiling Botanic Gardens (MBG)

The Mt. Makiling Forest Reserve (MMFR) stands as a highly biodiverse habitat and the only intact natural forest near Metro Manila, the Philippines. It is one of the 18 key centers of plant biodiversity and 32 key ecotourism sites in the Philippines. In monitoring the implementation plans for protecting MMFR, the information pertaining to the mushroom biodiversity across decades is important. Therefore, we aim to study mushroom as an indicator for biodiversity since there has been studies in the past 30 years on the macrofungi of MMFR which we summarized here along with ours. Sampling was done in August 2017 based on the transect line of 1000 m along roadsides of Makiling Botanic Gardens (MBG). The distribution of the sampling units was carried-out using random and stratified sampling. Our study describes 21 macrofungal taxa collected from MMFR. Of these, 20 taxa belong to Basidiomycota and only one belongs to Ascomycota. Polyporaceae was found as the most dominant macrofungi family (24%). There were six (6) species that are medicinal, and no poisonous species noted. There are eleven (11) species in this study which are unique records compared with previous studies done in the macrofungi of MMFR. This is the first study done comparing mushroom across 30 years on a reserved area. Information on these macrofungal flora across time serves as a reference for the currently existing conservation efforts and implementation of biodiversity-related policies in MMFR. © 2018, Society for Indonesian Biodiversity. All rights reserved

Preparation of Spin-Coated Poly(vinyl alcohol)/ chitosan/ Gold Nanoparticles Composite and Its Potential for Colorimetric Detection of Cyanide in Water

The rampant use of cyanide in mining activities has posed a substantial hazard to aquatic ecosystems and human health. Herein, a composite film of poly(vinyl alcohol)/chitosan (PVA/CS) with gold nanoparticles was synthesized as a potential colorimetric sensor for cyanide ions detection in water. The utilized AuNPs with a mean diameter of 14.32±3.49 nm were synthesized by chemical reduction using polyethyleneimine. Ultraviolet-visible spectroscopy revealed the characteristic peak of AuNPs at around 530 nm in the spectrum of PVA/CS/AuNP film, while SEM microscopy revealed the integration AuNPs across the polymer substrate. A color change from red to colorless was observed on the composite film after the addition of water samples with cyanide ions with a visual detection limit between 0.1-1 ppm. The sensor also exhibited good selectivity towards cyanide against the tested water samples containing various metal ions. This study reveals the potential of the fabricated PVA/CS/AuNP composite film as a simple, affordable, and environmentally benign material for on-site detection of cyanide in water and other related application

African army ants at the forefront of virome surveillance in a remote tropicalforest: A groundbreaking study using ants revealed a spectacular diversity of viruses in hardly accessible ecosystems like tropical forests

International audienceIn this study, we used a predator-enabled metagenomics strategy to sample the virome of a remote and difficult-to-access densely forested African tropical region. Specifically, we focused our study on the use of army ants of the genus Dorylus that are obligate collective foragers and group predators that attack and overwhelm a broad array of animal prey. Using 209 army ant samples collected from 29 colonies and the virion-associated nucleic acid-based metagenomics approach, we showed that a broad diversity of bacterial, plant, invertebrate and vertebrate viral sequences were accumulated by army ants: including sequences from 157 different viral genera in 56 viral families. This suggests that using predators and scavengers such as army ants to sample broad swathes of tropical forest viromes can shed light on the composition and the structure of viral populations of these complex and inaccessible ecosystems