A simple agar plating method for the description of microbial communities is described. This method is based on the quantification of the numbers of bacterial colonies in 6-7 age-based classes as they appear on agar media over a period of 6-10 days. The method can be used to quantify microbial communities in different habitats (roots and soil) and can be related to the ecophysiology of the microbial communities present. Significant differences in distribution patterns were found in time and depth on the roots. In general, as roots matured, the microbial communities changed from one dominated by r-strategists to one that was more distributed towards K-strategists. The soil had the greatest percentage of organisms that could be characterized as K-strategists. The method was also used to compare microbial communities on wheat roots and in soil in both the field and in microcosms in the glasshouse. In general, the method enabled differentiation between r- and K-strategists in environmental samples, something that could not be done using an ecophysiological index (a modification of the Shannon diversity index) or total bacterial numbers alone.
An isolate of Pseudomonas aureofaciens from the phylloplane of sugar beet which was chromosomally modified for monitors purposes by the insertion of two gene cassettes (kmr‐xylE and lacZY) was introduced to the phytosphere of spring wheat in a number of experiments and the resulting microbial perturbations quantified. Such studies involving innocuous bacterial isolates can serve as a guide in the assessment of risk associated with the release of functionally modified microorganisms. Introductions of P. aureofaciens on seeds caused large microbial perturbations (up to 2 log units) at the seedling stage on seeds and roots. As the inoculated plants matured (tillering, flowering and ripening), perturbations of total microbial populations were found to be non‐significant. Microbial perturbation on maturing wheat roots as a result of seed inoculations with P. aureofaciens could only be detected using more sensitive monitoring procedures describing the Pseudomonas community in terms of colony appearance rate on a selective Pseudomonas medium. Spray applications of the marked P. aureofaciens isolate onto the leaf surface of wheat caused no significant perturbations of the indigenous microbial present on the phylloplane.
Survival of Mycobacterium bovis after ingestion by protozoa would provide an environmental reservoir for infection of cattle. We have shown that M. bovis survived ingestion by Acanthamoeba castellanii. In contrast, two strains of M. bovis BCG did not survive well within Acanthamoeba.Bovine tuberculosis continues to pose an economic problem in Ireland and the United Kingdom. There is strong circumstantial evidence that badgers are a reservoir of the disease (14) and that ingestion of grass contaminated by badger waste is a likely route of infection. The length of survival of Mycobacterium bovis in the environment appears to vary considerably according to environmental conditions, ranging from only a few weeks (6, 24) to a year or more (18,26). However, there is scant understanding of how M. bovis survives in an environment to which it is ill adapted. Ingestion of M. bovis by protozoa might provide a protected environmental niche, enabling extended survival in the soil (4).Since the discovery that Legionella pneumophila can infect and replicate in amoebae (21), there has been increasing interest in the role of protozoa (especially Acanthamoeba spp.) in the environmental survival of pathogenic bacteria (4,7,8,16,17,23,25). Mycobacterium leprae and Mycobacterium avium have been shown to survive within protozoa (4,5,15,22), providing protection against adverse conditions (19). However, there is no published evidence that M. bovis can survive ingestion by amoebae.The purpose of this study was to test the survival of M. bovis in a model amoebal species, Acanthamoeba castellanii (Neff CCAP 1501/1A), which was maintained as axenic monolayers at 15°C in proteose-peptone-glucose medium (PPG). PPG is composed of 15 g of proteose-peptone, 18 g of D-glucose, and 1 liter of Page amoeba saline (PAS) (20). For bacterial infection, amoebal monolayers at approximately 90% confluency were resuspended, and viable counts of amoebae were determined by trypan blue exclusion in an improved Neubauer hemocytometer (11). Amoebae were added in 0.5 ml of PPG to each well of flat-bottom 24-well tissue culture plates (Nunc). For electron microscopy, monolayers were adhered to a piece of Melinex film in the tissue culture plate wells.After 2 h of incubation at 15°C, the PPG broth was removed and replaced with 0.5 ml of a bacterial suspension of M. bovis NCTC 10772 and two strains of M. bovis BCG (Pasteur, ATCC 35734; and Japan, laboratory collection), grown at 37°C to the log phase in Middlebrook 7H9 broth supplemented with 5% (vol/vol) OADC (oleic acid, albumin, dextrose, catalase enrichment; Becton Dickinson), 0.05% (vol/vol) Tween 80, and 4.1 g of sodium pyruvate per liter (for M. bovis) or 0.2% (vol/vol) glycerol (for BCG). The identity of the BCG strains was checked with IS6110 (9). Numbers of bacteria were estimated by optical density at 600 nm, and the cultures were resuspended in PAS for infection of Acanthamoeba at a multiplicity of infection of approximately 10, which was shown in preliminary experiments to achieve effective infection...
In a field release experiment, an isolate of Pseudomonas fluorescens, which was chromosomally modified with two reporter gene cassettes (lacZY and Kan r-xylE), was applied to spring wheat as a seed coating and subsequently as a foliar spray. The wild-type strain was isolated from the phylloplane of sugar beet but was found to be a common colonizer of both the rizosphere and phylloplane of wheat as well. The impact on the indigenous microbial populations resulting from release of this genetically modified microorganism (GMM) was compared with the impact of the unmodified, wild-type strain and a nontreated control until 1 month after harvest of the crop. The release of the P. fluorescens GMM and the unmodified, wild-type strain resulted in significant but transient perturbations of some of the culturable components of the indigenous microbial communities that inhabited the rhizosphere and phylloplane of wheat, but no significant perturbations of the indigenous culturable microbial populations in nonrhizosphere soil were found. Fast-growing organisms that did not produce resting structures (for example, fluorescent pseudomonads and yeasts) seemed to be most sensitive to perturbation. In terms of hazard and risk to the environment, the observed microbial perturbations that resulted from this GMM release may be considered minor for several reasons. First, the recombinant P. fluorescens strain caused changes that were, in general, not significantly different from those caused by the unmodified wild-type strain; second, perturbations resulting from bacterial inoculations were mainly small; and third, the release of bacteria had no obvious effects on plant growth and plant health.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.