Diversity of the spinach ( Spinacia oleracea ) spermosphere and phyllosphere bacterial communities

307

304

Seeds carry complex microbial communities, which may exert beneficial or deleterious effects on plant growth and plant health. To date, the composition of microbial communities associated with seeds has been explored mainly through culture-based diversity studies and therefore remains largely unknown. In this work, we analyzed the structures of the seed microbiotas of different plants from the family Brassicaceae and their dynamics during germination and emergence through sequencing of three molecular markers: the ITS1 region of the fungal internal transcribed spacer, the V4 region of 16S rRNA gene, and a species-specific bacterial marker based on a fragment of gyrB. Sequence analyses revealed important variations in microbial community composition between seed samples. Moreover, we found that emergence strongly influences the structure of the microbiota, with a marked reduction of bacterial and fungal diversity. This shift in the microbial community composition is mostly due to an increase in the relative abundance of some bacterial and fungal taxa possessing fast-growing abilities. Altogether, our results provide an estimation of the role of the seed as a source of inoculum for the seedling, which is crucial for practical applications in developing new strategies of inoculation for disease prevention.

Section: Discussionsupporting

confidence: 90%

mentioning

confidence: 99%

Emergence Shapes the Structure of the Seed Microbiota

Barret

Briand

Bonneau

et al. 2015

307

304

“…Similar to studies of other edible green vegetables, we found that the majority of bacteria on our leaf samples could be assigned to Pseudomonadales and Enterobacteriales (27,45,46). Although Methylobacterium spp.…”

Section: Discussionsupporting

confidence: 88%

High-Level Culturability of Epiphytic Bacteria and Frequency of Biosurfactant Producers on Leaves

Burch

Paulina

Sbodio

et al. 2016

To better characterize the bacterial community members capable of biosurfactant production on leaves, we distinguished culturable biosurfactant-producing bacteria from nonproducers and used community sequencing to compare the composition of these distinct cultured populations with that from DNA directly recovered from leaves. Communities on spinach, romaine, and head lettuce leaves were compared with communities from adjacent samples of soil and irrigation source water. Soil communities were poorly described by culturing, with recovery of cultured representatives from only 21% of the prevalent operational taxonomic units (OTUs) (>0.2% reads) identified. The dominant biosurfactant producers cultured from soil included bacilli and pseudomonads. In contrast, the cultured communities from leaves are highly representative of the culture-independent communities, with over 85% of the prevalent OTUs recovered. The dominant taxa of surfactant producers from leaves were pseudomonads as well as members of the infrequently studied genus Chryseobacterium. The proportions of bacteria cultured from head lettuce and romaine leaves that produce biosurfactants were directly correlated with the culture-independent proportion of pseudomonads in a given sample, whereas spinach harbored a wider diversity of biosurfactant producers. A subset of the culturable bacteria in irrigation water also became enriched on romaine leaves that were irrigated overhead. Although our study was designed to identify surfactant producers on plants, we also provide evidence that most bacteria in some habitats, such as agronomic plant surfaces, are culturable, and these communities can be readily investigated and described by more classical culturing methods. IMPORTANCEThe importance of biosurfactant production to the bacteria that live on waxy leaf surfaces as well as their ability to be accurately assessed using culture-based methodologies was determined by interrogating epiphytic populations by both culture-dependent and culture-independent methods. Biosurfactant production was much more frequently observed in cultured communities on leaves than in other nearby habitats, such as soil and water, suggesting that this trait is important to life on a leaf by altering either the leaf itself or the interaction of bacteria with water. While pseudomonads were the most common biosurfactant producers isolated, this habitat also selects for taxa, such as Chryseobacterium, for which this trait was previously unrecognized. The finding that most epiphytic bacterial taxa were culturable validates strategies using more classical culturing methodologies for their study in this habitat. The phyllosphere is recognized to be a selective habitat in which epiphytic bacteria must be able to access limited and spatially heterogeneous nutrient supplies and endure frequent fluctuations in moisture availability on a water-repellent surface (1, 2). Recent culture-independent community analyses have confirmed the apparent selectivity of leaf surfaces, finding that leaf sur...

“…In addition, the short period of stress application (2 days in this study) might not have significant effects on bacterial counts, which are known to change over the long growing season (65,66). On the other hand, the structure of phyllosphere bacterial communities can be influenced by a number of environmental factors (63,67,68), which may explain the various significant correlations between the different viruses and the bacterial counts on lettuce and spinach observed under different conditions. In our study, both the levels of bacteria and the virus infectivity and RNA titers showed significant positive correlations under multiple conditions for multiple viruses.…”

Section: Discussionmentioning

confidence: 83%

Abiotic Stress and Phyllosphere Bacteria Influence the Survival of Human Norovirus and Its Surrogates on Preharvest Leafy Greens

Esseili

Gao

Tegtmeier

et al. 2016

Foodborne outbreaks of human noroviruses (HuNoVs) are frequently associated with leafy greens. Because there is no effective method to eliminate HuNoV from postharvest leafy greens, understanding virus survival under preharvest conditions is crucial. The objective of this study was to evaluate the survival of HuNoV and its surrogate viruses, murine norovirus (MNV), porcine sapovirus (SaV), and Tulane virus (TV), on preharvest lettuce and spinach that were subjected to abiotic stress (physical damage, heat, or flood). We also examined the bacteria culturable from the phyllosphere in response to abiotic stress and in relation to viral persistence. Mature plants were subjected to stressors 2 days prior to inoculation of the viruses on leaves. We quantified the viral RNA, determined the infectivity of the surrogates, and performed bacterial counts on postinoculation days (PIDs) 0, 1, 7, and 14. For both plant types, time exerted significant effects on HuNoV, MNV, SaV, and TV RNA titers, with greater effects being seen for the surrogates. Infectious surrogate viruses were undetectable on PID 14. Only physical damage on PID 14 significantly enhanced HuNoV RNA persistence on lettuce, while the three stressors differentially enhanced the persistence of MNV and TV RNA. Bacterial counts were significantly affected by time and plant type but not by the stressors. However, bacterial counts correlated significantly with HuNoV RNA titers on spinach and with the presence of surrogate viruses on both plant types under various conditions. In conclusion, abiotic stressors and phyllosphere bacterial density may differentially influence the survival of HuNoV and its surrogates on lettuce and spinach, emphasizing the need for the use of preventive measures at the preharvest stage.H uman noroviruses (HuNoVs) are the leading cause of acute gastroenteritis and foodborne illnesses in the United States, causing 19 million to 21 million cases, 570 to 800 deaths, and $777 million in health care-associated costs annually (1). These singlestranded RNA viruses belong to the Caliciviridae family and are 28 to 35 nm in diameter and nonenveloped. Although these viruses cause self-limiting gastroenteritis, more serious infections can develop in high-risk groups, such as the elderly and immunocompromised populations (2). The virus is mainly transmitted to humans via the fecal-oral route either by contact with an infected person or fomites or by ingestion of contaminated food and water (2). Leafy greens are frequently associated with HuNoV outbreaks (1) and are globally recognized to be a high priority in terms of the microbial safety of fresh produce (3). Contamination of leafy greens with HuNoVs can occur at any stage along the farm-to-fork chain (3) through a number of sources, including fecally contaminated water used for irrigation, improperly treated sewage sludge used for fertilization, and food harvesters or food handlers who use improper hygiene practices (4, 5). Because HuNoVs have a low infectious dose (ϳ18 to 1,000 viral particles) (6) ...