Freshwater plastispheres as a vector for foodborne bacteria and viruses

There is growing evidence that plastic particles can accumulate microorganisms that are pathogenic to humans or animals. In the current study, the composition of the plastispheres that accumulated on polypropylene (PP), polyvinyl chloride (PVC), and high-density polyethylene (HDPE) pieces submerged in a river in the southeast Norway was characterized by 16S rRNA amplicon sequencing. Seasonal and geographical effects on the bacterial composition of the plastisphere were identified, in addition to the detection of potential foodborne pathogenic bacteria and viruses as part of the plastisphere. The diversity and taxonomic composition of the plastispheres were influenced by the number of weeks in the river, the season, and the location. The bacterial diversity differed significantly in the plastisphere from June and September, with a generally higher diversity in June. Also, the community composition of the plastisphere was significantly influenced by the geographical location, while the type of plastic had less impact. Plastics submerged in river water assembled a variety of microorganisms including potentially pathogenic bacteria and viruses (noro- and adenovirus) detected by qPCR. Cultivation methods detected viable bacteria such as Escherichia coli and Listeria monocytogenes. The results highlight the need for additional research on the risk of contaminating food with plastic particles colonized with human pathogens through irrigation water.     

Distinct spatiotemporal succession of bacterial generalists and specialists in the lacustrine plastisphere

The assembly processes of generalists and specialists and their driving mechanisms during spatiotemporal succession is a central issue in microbial ecology but a poorly researched subject in the plastisphere. We investigated the composition variation, spatiotemporal succession, and assembly processes of bacterial generalists and specialists in the plastisphere, including non-biodegradable (NBMPs) and biodegradable microplastics (BMPs). Although the composition of generalists and specialists on NBMPs differed from that of BMPs, colonization time mainly mediated the composition variation. The relative abundance of generalists and the relative contribution of species replacement were initially increased and then decreased with colonization time, while the specialists initially decreased and then increased. Besides, the richness differences also affected the composition variation of generalists and specialists in the plastisphere, and the generalists were more susceptible to richness differences than corresponding specialists. Furthermore, the assembly of generalists in the plastisphere was dominated by deterministic processes, while stochastic processes dominated the assembly of specialists. The network stability test showed that the community stability of generalists on NBMPs and BMPs was lower than corresponding specialists. Our results suggested that different ecological assembly processes shaped the spatiotemporal succession of bacterial generalists and specialists in the plastisphere, but were less influenced by polymer types.     

Contribution of stochastic processes to the microbial community assembly on field-collected microplastics

A growing body of evidence suggests that microplastics may be colonized with a unique microbiome, termed ‘plastisphere’, in aquatic environments. However, the deep mechanisms (deterministic and/or stochastic processes) underlying the community assembly on microplastics are still poorly understood. Here, we took the estuary of Hangzhou Bay (Zhejiang, China) as an example and examined the assembly mechanisms of bacterial communities in water and microplastic samples. Results from high-throughput sequencing showed that Proteobacteria, Firmicutes, and Actinobacteria were the dominant phyla across all samples. Additionally, microorganisms from plastisphere and planktonic communities exhibited contrasting taxonomic compositions, with greater within-group variation for microplastic samples. The null model analysis indicated the plastisphere bacterial communities were dominantly driven by the stochastic process of drift (58.34%) and dispersal limitation (23.41%). The normalized stochasticity ratio (NST) also showed that the community assembly on microplastics was more stochastic (NST > 50%). Based on the Sloan neutral community model, the migration rate for plastisphere communities (0.015) was significantly lower than that for planktonic communities (0.936), potentially suggesting that it is the stochastic balance between loss and gain of bacteria (e.g., stochastic births and deaths) critically shaping the community assembly on microplastics and generating the specific niches. This study greatly enhanced our understanding of the ecological patterns of microplastic-associated microbial communities in aquatic environments.     

Short-term plastisphere colonization dynamics across six plastic types

Marine plastic pollution is a major concern worldwide, but the understanding of plastisphere dynamics remains limited in the southern hemisphere. To address this knowledge gap, we conducted a study in South Australia to investigate the prokaryotic community of the plastisphere and its temporal changes over 4 weeks. We submerged six plastic types (i.e., High-Density Polyethylene [HDPE], Polyvinyl chloride [PVC], Low-Density Polyethylene [LDPE], Polypropylene [PP], Polystyrene [PS] and the understudied textile, polyester [PET]) and wood in seawater and sampled them weekly to characterize the prokaryotic community using 16S rRNA gene metabarcoding. Our results showed that the plastisphere composition shifted significantly over short time scales (i.e., 4 weeks), and each plastic type had distinct groups of unique genera. In particular, the PVC plastisphere was dominated by Cellvibrionaceae taxa, distinguishing it from other plastics. Additionally, the textile polyester, which is rarely studied in plastisphere research, supported the growth of a unique group of 25 prokaryotic genera (which included the potential pathogenic Legionella genus). Overall, this study provides valuable insights into the colonization dynamics of the plastisphere over short time scales and contributes to narrowing the research gap on the southern hemisphere plastisphere.     

Soil plastispheres as hotspots of antibiotic resistance genes and potential pathogens

In the Anthropocene, increasing pervasive plastic pollution is creating a new environmental compartment, the plastisphere. How the plastisphere affects microbial communities and antibiotic resistance genes (ARGs) is an issue of global concern. Although this has been studied in aquatic ecosystems, our understanding of plastisphere microbiota in soil ecosystems remains poor. Here, we investigated plastisphere microbiota and ARGs of four types of microplastics (MPs) from diverse soil environments, and revealed effects of manure, temperature, and moisture on them. Our results showed that the MPs select for microbial communities in the plastisphere, and that these plastisphere communities are involved in diverse metabolic pathways, indicating that they could drive diverse ecological processes in the soil ecosystem. The relationship within plastisphere bacterial zero-radius operational taxonomic units (zOTUs) was predominantly positive, and neutral processes appeared to dominate community assembly. However, deterministic processes were more important in explaining the variance in ARGs in plastispheres. A range of potential pathogens and ARGs were detected in the plastisphere, which were enriched compared to the soil but varied across MPs and soil types. We further found that the addition of manure and elevation of soil temperature and moisture all enhance ARGs in plastispheres, and potential pathogens increase with soil moisture. These results suggested that plastispheres are habitats in which an increased potential pathogen abundance is spatially co-located with an increased abundance of ARGs under global change. Our findings provided new insights into the community ecology of the microbiome and antibiotic resistome of the soil plastisphere.Subject terms: Microbial ecology, Environmental sciences     

Microplastic polymer properties as deterministic factors driving terrestrial plastisphere microbiome assembly and succession in the field

Environmental microplastic (MP) is ubiquitous in aquatic and terrestrial ecosystems providing artificial habitats for microbes. Mechanisms of MP colonization, MP polymer impacts, and effects on soil microbiomes are largely unknown in terrestrial systems. Therefore, we experimentally tested the hypothesis that MP polymer type is an important deterministic factor affecting MP community assembly by incubating common MP polymer types in situ in landfill soil for 14 months. 16S rRNA gene amplicon sequencing indicated that MP polymers have specific impacts on plastisphere microbiomes, which are subsets of the soil microbiome. Chloroflexota, Gammaproteobacteria, certain Nitrososphaerota, and Nanoarchaeota explained differences among MP polymers and time points. Plastisphere microbial community composition derived from different MP diverged over time and was enriched in potential pathogens. PICRUSt predictions of pathway abundances and quantitative PCR of functional marker genes indicated that MP polymers exerted an ambivalent effect on genetic potentials of biogeochemical cycles. Overall, the data indicate that (i) polymer type as deterministic factor rather than stochastic factors drives plastisphere community assembly, (ii) MP impacts greenhouse gas metabolism, xenobiotic degradation and pathogen distribution, and (iii) MP serves as an ideal model system for studying fundamental questions in microbial ecology such as community assembly mechanisms in terrestrial environments.     

Pyrosequencing analysis of bacterial community and assembly in activated sludge samples from different geographic regions in China

In order to investigate if there are geographic differences of bacterial community in the activated sludge collected from different geographic regions (eastern, northwestern, and northern parts) of China and to determine the co-occurrence patterns of bacterial community, activated sludge samples were collected from 10 municipal wastewater treatment plants located in 8 cities in China. High-throughput pyrosequencing combined with the bioinformatics analysis were used to examine the bacterial community compositions in the activated sludge samples. The result of taxonomy classifier indicated that a total of 76 genera were commonly shared by more than 7 samples, which accounted for 62 to 96 % of the classified sequences in each sample. Even though some core genera existed in all examined activated sludge samples regardless of the sampling geographic location and treatment process, significant geographic differences of bacterial community compositions among the activated sludge samples were revealed by the nonmetric multidimensional scaling (NMDS) and analyses of similarity (ANOSIM) analysis. A total of 165 pairs of significant and robust correlations (positive and negative) were identified from 61 bacterial genera based on the network analysis. The data obtained in this study could provide useful information to understand the bacterial community composition in geographically distributed wastewater treatment plants and discern the co-occurrence patterns of bacterial community.     

Litter quality versus soil microbial community controls over decomposition: a quantitative analysis

The possible effects of soil microbial community structure on organic matter decomposition rates have been widely acknowledged, but are poorly understood. Understanding these relationships is complicated by the fact that microbial community structure and function are likely to both affect and be affected by organic matter quality and chemistry, thus it is difficult to draw mechanistic conclusions from field studies. We conducted a reciprocal soil inoculum × litter transplant laboratory incubation experiment using samples collected from a set of sites that have similar climate and plant species composition but vary significantly in bacterial community structure and litter quality. The results showed that litter quality explained the majority of variation in decomposition rates under controlled laboratory conditions: over the course of the 162-day incubation, litter quality explained nearly two-thirds (64 %) of variation in decomposition rates, and a smaller proportion (25 %) was explained by variation in the inoculum type. In addition, the relative importance of inoculum type on soil respiration increased over the course of the experiment, and was significantly higher in microcosms with lower litter quality relative to those with higher quality litter. We also used molecular phylogenetics to examine the relationships between bacterial community composition and soil respiration in samples through time. Pyrosequencing revealed that bacterial community composition explained 32 % of the variation in respiration rates. However, equal portions (i.e., 16 %) of the variation in bacterial community composition were explained by inoculum type and litter quality, reflecting the importance of both the meta-community and the environment in bacterial assembly. Taken together, these results indicate that the effects of changing microbial community composition on decomposition are likely to be smaller than the potential effects of climate change and/or litter quality changes in response to increasing atmospheric CO₂ concentrations or atmospheric nutrient deposition.     

Microplastics reduce soil microbial network complexity and ecological deterministic selection

Microplastics have been proposed as emerging threats for terrestrial systems as they may potentially alter the physicochemical/biophysical soil environments. Due to the variety of properties of microplastics and soils, the microplastic-induced effects in soil ecosystems are greatly manifold. Here, we studied effects of three polymer microplastics (polyamide-6, polyethylene, and polyethylene terephthalate) on soil properties with four different soil types. The success patterns, interaction relationships, and assembly processes of soil bacterial communities were also studied. Microplastics have the potential to promote CO₂ emissions and enhance the soil humification. Even though microplastics did not significantly alter the diversity and composition of the soil microbial community, the application of microplastics decreased the network complexity and stability, including network size, connectivity, and the number of module and keystone species. The bacterial community assembly was governed by deterministic selection (77.3%–90.9%) in all treatments, while microplastics increased the contribution of stochastic processes from 9.1% in control to 13.6%–22.7%. The neutral model results also indicated most of the bacterial taxa were present in the predicted neutral region (approximately 98%), suggesting the importance of stochastic processes. These findings provided a fundamental insight in understanding the effects of microplastics on soil ecosystems.     

Microplastics as a New Ecological Niche For Multispecies Microbial Biofilms within the Plastisphere

Microbiology - The distribution, colonization, and biodegradation of primary and secondary microplastics, as well as were effect on the plastisphere ecological systems, are analyzed. The pathways...     

Different biogeographic patterns of prokaryotes and microbial eukaryotes in epilithic biofilms

Microbial biogeography studies expend much effort in determining whether environmental selection or stochastic processes related to dispersal are more important in shaping community composition. While both types of factors are possibly influential, it is tacitly assumed that protists, or microbial eukaryotes in general, behave biogeographically as prokaryotes because of their small physical size. However, direct evidence for this in exactly the same environment and at the same phylogenetic depth is lacking. In this study, we compared the structure of both prokaryotic and eukaryotic components of microbial communities forming biofilms on mineral substrates in different geographic locations at the level of small-subunit (SSU) rRNA-based operational taxonomic units (OTUs). These microbial communities are subjected to strong environmental selection and contain significant proportions of extremophilic microorganisms adapted to desiccation and UV radiation. We find that the nature of the substrate as well as climatic variables and geography influences microbial community structure. However, constrained correspondence analyses and distance-decay curves showed that, whereas the substrate type was the most significant factor structuring bacterial communities, geographic location was the most influential factor for microbial eukaryote communities. Biological explanations implying a higher dispersal success for bacteria combined with more mobile lifestyles for predatory protists may underlie these different prokaryote versus microbial eukaryote biogeographic patterns.     

Analysis of bacterial and archaeal diversity in coastal microbial mats using massive parallel 16S rRNA gene tag sequencing

Coastal microbial mats are small-scale and largely closed ecosystems in which a plethora of different functional groups of microorganisms are responsible for the biogeochemical cycling of the elements. Coastal microbial mats play an important role in coastal protection and morphodynamics through stabilization of the sediments and by initiating the development of salt-marshes. Little is known about the bacterial and especially archaeal diversity and how it contributes to the ecological functioning of coastal microbial mats. Here, we analyzed three different types of coastal microbial mats that are located along a tidal gradient and can be characterized as marine (ST2), brackish (ST3) and freshwater (ST3) systems. The mats were sampled during three different seasons and subjected to massive parallel tag sequencing of the V6 region of the 16S rRNA genes of Bacteria and Archaea. Sequence analysis revealed that the mats are among the most diverse marine ecosystems studied so far and consist of several novel taxonomic levels ranging from classes to species. The diversity between the different mat types was far more pronounced than the changes between the different seasons at one location. The archaeal community for these mats have not been studied before and revealed a strong reaction on a short period of draught during summer resulting in a massive increase in halobacterial sequences, whereas the bacterial community was barely affected. We concluded that the community composition and the microbial diversity were intrinsic of the mat type and depend on the location along the tidal gradient indicating a relation with salinity.     

Jellyfish modulate bacterial dynamic and community structure

Jellyfish blooms have increased in coastal areas around the world and the outbreaks have become longer and more frequent over the past few decades. The Mediterranean Sea is among the heavily affected regions and the common bloom-forming taxa are scyphozoans Aurelia aurita s.l., Pelagia noctiluca, and Rhizostoma pulmo. Jellyfish have few natural predators, therefore their carcasses at the termination of a bloom represent an organic-rich substrate that supports rapid bacterial growth, and may have a large impact on the surrounding environment. The focus of this study was to explore whether jellyfish substrate have an impact on bacterial community phylotype selection. We conducted in situ jellyfish-enrichment experiment with three different jellyfish species. Bacterial dynamic together with nutrients were monitored to assess decaying jellyfish-bacteria dynamics. Our results show that jellyfish biomass is characterized by protein rich organic matter, which is highly bioavailable to 'jellyfish-associated' and 'free-living' bacteria, and triggers rapid shifts in bacterial population dynamics and composition. Based on 16S rRNA clone libraries and denaturing gradient gel electrophoresis (DGGE) analysis, we observed a rapid shift in community composition from unculturable Alphaproteobacteria to culturable species of Gammaproteobacteria and Flavobacteria. The results of sequence analyses of bacterial isolates and of total bacterial community determined by culture independent genetic analysis showed the dominance of the Pseudoalteromonadaceae and the Vibrionaceae families. Elevated levels of dissolved proteins, dissolved organic and inorganic nutrient release, bacterial abundance and carbon production as well as ammonium concentrations characterized the degradation process. The biochemical composition of jellyfish species may influence changes in the amount of accumulated dissolved organic and inorganic nutrients. Our results can contribute insights into possible changes in bacterial population dynamics and nutrient pathways following jellyfish blooms which have important implications for ecology of coastal waters.     

Effects of disturbance intensity and frequency on bacterial community composition and function

Disturbances influence community structure and ecosystem functioning. Bacteria are key players in ecosystems and it is therefore crucial to understand the effect of disturbances on bacterial communities and how they respond to them, both compositionally and functionally. The main aim of this study was to test the effect of differences in disturbance strength on bacterial communities. For this, we implemented two independent short-term experiments with dialysis bags containing natural bacterial communities, which were transplanted between ambient and 'disturbed' incubation tanks, manipulating either the intensity or the frequency of a salinity disturbance. We followed changes in community composition by terminal restriction fragment analysis (T-RFLP) and measured various community functions (bacterial production, carbon substrate utilization profiles and rates) directly after and after a short period of recovery under ambient conditions. Increases in disturbance strength resulted in gradually stronger changes in bacterial community composition and functions. In the disturbance intensity experiment, the sensitivity to the disturbance and the ability of recovery differed between different functions. In the disturbance frequency experiment, effects on the different functions were more consistent and recovery was not observed. Moreover, in case of the intensity experiment, there was also a time lag in the responses of community composition and functions, with functional responses being faster than compositional ones. To summarize, our study shows that disturbance strength has the potential to change the functional performance and composition of bacterial communities. It further highlights that the overall effects, rates of recovery and the degree of congruence in the response patterns of community composition and functioning along disturbance gradients depend on the type of function and the character of the disturbance.     

Temperature controls on aquatic bacterial production and community dynamics in arctic lakes and streams

The impact of temperature on bacterial activity and community composition was investigated in arctic lakes and streams in northern Alaska. Aquatic bacterial communities incubated at different temperatures had different rates of production, as measured by ¹⁴C‐leucine uptake, indicating that populations within the communities had different temperature optima. Samples from Toolik Lake inlet and outlet were collected at water temperatures of 14.2°C and 15.9°C, respectively, and subsamples incubated at temperatures ranging from 6°C to 20°C. After 5 days, productivity rates varied from 0.5 to ～13.7 µg C l^?1 day^?1 and two distinct activity optima appeared at 12°C and 20°C. At these optima, activity was 2‐ to 11‐fold higher than at other incubation temperatures. The presence of two temperature optima indicates psychrophilic and psychrotolerant bacteria dominate under different conditions. Community fingerprinting via denaturant gradient gel electrophoresis (DGGE) of 16S rRNA genes showed strong shifts in the composition of communities driven more by temperature than by differences in dissolved organic matter source; e.g. four and seven unique operational taxonomic units (OTUs) were found only at 2°C and 25°C, respectively, and not found at other incubation temperatures after 5 days. The impact of temperature on bacteria is complex, influencing both bacterial productivity and community composition. Path analysis of measurements of 24 streams and lakes sampled across a catchment 12 times in 4 years indicates variable timing and strength of correlation between temperature and bacterial production, possibly due to bacterial community differences between sites. As indicated by both field and laboratory experiments, shifts in dominant community members can occur on ecologically relevant time scales (days), and have important implications for understanding the relationship of bacterial diversity and function.     

Availability of glucose and light modulates the structure and function of a microbial biofilm

We have studied the differences in the organic matter processing and biofilm composition and structure between autoheterotrophic and heterotrophic biofilm communities. Microbial communities grown on artificial biofilms were monitored, following incubation under light and dark conditions and with or without the addition of glucose as a labile organic compound. Glucose addition greatly affected the microbial biofilm composition as shown by differences in 16S rRNA gene fingerprints. A significant increase in β-glucosidase and peptidase enzyme activities were also observed in glucose-amended biofilms incubated in the dark, suggesting an active bacterial community. Light enhanced the algal and bacterial growth, as well as higher extracellular enzyme activity, thereby indicating a tight algal–bacterial coupling in biofilms incubated under illumination. In these biofilms, organic compounds excreted by photosynthetic microorganisms were readily available for bacterial heterotrophs. This algal–bacterial relationship weakened in glucose-amended biofilms grown in the light, probably because heterotrophic bacteria preferentially use external labile compounds. These results suggest that the availability of labile organic matter in the flowing water and the presence of light may alter the biofilm composition and function, therefore affecting the processing capacity of organic matter in the stream ecosystem.     

The ecology of the phyllosphere: geographic and phylogenetic variability in the distribution of bacteria on tree leaves

Large populations of bacteria live on leaf surfaces and these phyllosphere bacteria can have important effects on plant health. However, we currently have a limited understanding of bacterial diversity on tree leaves and the inter‐ and intra‐specific variability in phyllosphere community structure. We used a barcoded pyrosequencing technique to characterize the bacterial communities from leaves of 56 tree species in Boulder, Colorado, USA, quantifying the intra‐ and inter‐individual variability in the bacterial communities from 10 of these species. We also examined the geographic variability in phyllosphere communities on Pinus ponderosa from several locations across the globe. Individual tree species harboured high levels of bacterial diversity and there was considerable variability in community composition between trees. The bacterial communities were organized in patterns predictable from the relatedness of the trees as there was significant correspondence between tree phylogeny and bacterial community phylogeny. Inter‐specific variability in bacterial community composition exceeded intra‐specific variability, a pattern that held even across continents where we observed minimal geographic differentiation in the bacterial communities on P. ponderosa needles.     

Microbes and microplastics: Community shifts along an urban coastal contaminant gradient

Plastic substrates introduced to the environment during the Anthropocene have introduced new pathways for microbial selection and dispersal. Some plastic-colonising microorganisms have adapted phenotypes for plastic degradation (selection), while the spatial transport (dispersal) potential of plastic colonisers remains controlled by polymer-specific density, hydrography and currents. Plastic-degrading enzyme abundances have recently been correlated with concentrations of plastic debris in open ocean environments, making it critical to better understand colonisation of hydrocarbon degraders with plastic degradation potential in urbanised watersheds where plastic pollution often originates. We found that microbial colonisation by reputed hydrocarbon degraders on microplastics (MPs) correlated with a spatial contaminant gradient (New York City/Long Island waterways), polymer types, temporal scales, microbial domains and putative cell activity (DNA vs. RNA). Hydrocarbon-degrading taxa enriched on polyethylene and polyvinyl chloride substrates relative to other polymers and were more commonly recovered in samples proximal to New York City. These differences in MP colonisation could indicate phenotypic adaptation processes resulting from increased exposure to urban plastic runoff as well as differences in carbon bioavailability across polymer types. Shifts in MP community potential across urban coastal contaminant gradients and polymer types improve our understanding of environmental plastic discharge impacts toward biogeochemical cycling across the global ocean.     

Pyrosequencing analysis of bacterial diversity in 14 wastewater treatment systems in china

To determine if there is a core microbial community in the microbial populations of different wastewater treatment plants (WWTPs) and to investigate the effects of wastewater characteristics, operational parameters, and geographic locations on microbial communities, activated sludge samples were collected from 14 wastewater treatment systems located in 4 cities in China. High-throughput pyrosequencing was used to examine the 16S rRNA genes of bacteria in the wastewater treatment systems. Our results showed that there were 60 genera of bacterial populations commonly shared by all 14 samples, including Ferruginibacter, Prosthecobacter, Zoogloea, Subdivision 3 genera incertae sedis, Gp4, Gp6, etc., indicating that there is a core microbial community in the microbial populations of WWTPs at different geographic locations. The canonical correspondence analysis (CCA) results showed that the bacterial community variance correlated most strongly with water temperature, conductivity, pH, and dissolved oxygen (DO) content. Variance partitioning analyses suggested that wastewater characteristics had the greatest contribution to the bacterial community variance, explaining 25.7% of the variance of bacterial communities independently, followed by operational parameters (23.9%) and geographic location (14.7%). Results of this study provided insights into the bacterial community structure and diversity in geographically distributed WWTPs and discerned the relationships between bacterial community and environmental variables in WWTPs.     

Effect of Incubation on Bacterial Communities of Eggshells in a Temperate Bird,the Eurasian Magpie (Pica pica)

Inhibitory effect of incubation on microbial growth has extensively been studied in wild bird populations using culture-based methods and conflicting results exist on whether incubation selectively affects the growth of microbes on the egg surface. In this study, we employed culture-independent methods, quantitative PCR and 16S rRNA gene pyrosequencing, to elucidate the effect of incubation on the bacterial abundance and bacterial community composition on the eggshells of the Eurasian Magpie (Pica pica). We found that total bacterial abundance increased and diversity decreased on incubated eggs while there were no changes on non-incubated eggs. Interestingly, Gram-positive Bacillus, which include mostly harmless species, became dominant and genus Pseudomonas, which include opportunistic avian egg pathogens, were significantly reduced after incubation. These results suggest that avian incubation in temperate regions may promote the growth of harmless (or benevolent) bacteria and suppress the growth of pathogenic bacterial taxa and consequently reduce the diversity of microbes on the egg surface. We hypothesize that this may occur due to difference in sensitivity to dehydration on the egg surface among microbes, combined with the introduction of Bacillus from bird feathers and due to the presence of antibiotics that certain bacteria produce.