RESUMEN
Measuring the abundance of microbes in a sample is a common procedure with a long history, but best practices are not well-conserved across microbiological ï¬elds. Serial dilution methods are commonly used to dilute bacterial cultures to produce countable numbers of colonies, and from these counts, to infer bacterial concentrations measured in colony-forming units (CFUs). The most common methods to generate data for CFU point estimates involve plating bacteria on (or in) a solid growth medium and counting their resulting colonies or counting the number of tubes at a given dilution that have growth. Traditionally, these types of data have been analyzed separately using different analytic methods. Here, we build a direct correspondence between these approaches, which allows one to extend the use of the most probable number method from the liquid tubes experiments, for which it was developed, to the growth plates by viewing colony-sized patches of a plate as equivalent to individual tubes. We also discuss how to combine measurements taken at different dilutions, and we review several ways of analyzing colony counts, including the Poisson and truncated Poisson methods. We test all point estimate methods computationally using simulated data. For all methods, we discuss their relevant error bounds, assumptions, strengths, and weaknesses. We provide an online calculator for these estimators.Estimation of the number of microbes in a sample is an important problem with a long history. Yet common practices, such as combining results from different measurements, remain sub-optimal. We provide a comparison of methods for estimating abundance of microbes and detail a mapping between different methods, which allows to extend their range of applicability. This mapping enables higher precision estimates of colony-forming units (CFUs) using the same data already collected for traditional CFU estimation methods. Furthermore, we provide recommendations for how to combine measurements of colony counts taken across dilutions, correcting several misconceptions in the literature.
Asunto(s)
Bacterias , Recuento de Colonia Microbiana , Bacterias/aislamiento & purificación , Bacterias/clasificación , Bacterias/crecimiento & desarrollo , Recuento de Colonia Microbiana/métodos , Funciones de Verosimilitud , Distribución de PoissonRESUMEN
Trade-offs between competitive ability and resistance to top-down control manifest the "kill-the-winner" hypothesis that explains how mortality caused by protists and viruses can promote bacterial diversity. However, the existence of such trade-offs has rarely been investigated in natural marine bacterial communities. To address this question, we conducted on-board dilution experiments to manipulate top-down control pressure (protists only or protists plus viruses [protists+viruses] combined) and then applied 16S rRNA gene high-throughput sequencing techniques to assess the responses of each bacterial taxon. Dilution experiments enabled us to measure the top-down-control-free growth rate as the competitive ability and top-down-control-caused mortality as the reverse of resistance to top-down control. Overall, bacterial taxa with higher top-down-control-free growth rates were accompanied by lower top-down-control-caused resistance. Furthermore, competition-resistance trade-offs were stronger and more consistent when top-down control was caused by protists+viruses combined than by protists only. When protists+viruses were diluted, the bacterial rank abundance distribution became steepened and evenness and richness were decreased. However, when protists were diluted, only richness decreased. Our results indicate the existence of competition-resistance trade-offs in marine microbes and demonstrate the positive impacts of such trade-offs on bacterial diversity. Regardless, the strength of the competition-resistance trade-offs and the impacts on bacterial diversity were contingent on whether top-down control was caused by protists+viruses combined or protists only. IMPORTANCE We addressed the "kill-the-winner" hypothesis from the perspective of its principle (the competition-resistance trade-off) in marine bacterial communities incubated in situ. Our results supported the existence of competition-resistance trade-offs and the positive effect on bacterial community diversity. The study linked theoretical expectations and complex natural systems and provided new knowledge regarding how top-down controls and competition trade-offs shaped natural bacterial communities.
Asunto(s)
Virus , ARN Ribosómico 16S/genética , Bacterias/genéticaRESUMEN
Microzooplankton are considered the primary consumers of phytoplankton in marine environments. Microzooplankton grazing rates on phytoplankton have been studied across the globe, but there are still large regions of the ocean that are understudied, such as sub-tropical coastal oceans. One of these regions is the coastal area around south Florida, USA. We measured microzooplankton grazing rates in two distinct environments around south Florida; the oligotrophic Florida Keys and the mesotrophic outflow from the Everglades. For 2-years from January 2018 to January 2020, we set up 55 dilution and light-dark bottle experiments at five stations to estimate the microzooplankton community grazing rate, instantaneous phytoplankton growth rate, and primary production. Our results suggest that microzooplankton are consuming a higher proportion of the primary production near the Everglades outflow compared to the Florida Keys. We also found that changes in phytoplankton growth rates are disconnected from changes in the microzooplankton grazing rates in the Florida Keys. Overall, the data from the Everglades outflow is what would be expected based on global patterns, but factors other than microzooplankton grazing are more important in controlling phytoplankton biomass in the Florida Keys.
Asunto(s)
Fitoplancton , Florida , Océanos y Mares , BiomasaRESUMEN
Behaviors of dynamic polymers such as microtubules and actin are frequently assessed at one or both of the following scales: (a) net assembly or disassembly of bulk polymer, (b) growth and shortening of individual filaments. Previous work has derived various forms of an equation to relate the rate of change in bulk polymer mass (i.e., flux of subunits into and out of polymer, often abbreviated as "J") to individual filament behaviors. However, these versions of the "J equation" differ in the variables used to quantify individual filament behavior, which correspond to different experimental approaches. For example, some variants of the J equation use dynamic instability parameters, obtained by following particular individual filaments for long periods of time. Another form of the equation uses measurements from many individuals followed over short time steps. We use a combination of derivations and computer simulations that mimic experiments to (a) relate the various forms of the J equation to each other, (b) determine conditions under which these J equation forms are and are not equivalent, and (c) identify aspects of the measurements that can affect the accuracy of each form of the J equation. Improved understanding of the J equation and its connections to experimentally measurable quantities will contribute to efforts to build a multiscale understanding of steady-state polymer behavior.