Role In Germination Research Articles

Germination and growth responses to water stress of three agroforestry tree species from Bangladesh

Globally, tree plantations are considered one of the most effective solutions in tackling the climate crisis, and by incorporating trees, agroforestry plays an essential role in mitigating climate change. However, the productivity and sustainability of this commendable solution are often impeded by the wide range of abiotic stresses, including drought. Species selection is always considered as one of the major challenges in a drought-prone area to ensure maximum productivity and halting tree mortality. In this study, seedling responses such as germination, survival, and growth were examined for Swietenia macrophylla, Acacia nilotica and Pithecellobium dulce in variable soil moisture regimes with 100%, 50% and 25% soil water content (SWC). Despite complete failure of germination at 25% SWC, S. macrophylla showed a significantly higher germination percent at both 100% and 50% SWCs (p < 0.05). Irrespective of species, the 25% SWC showed reductions of germination percent, survivability and total dry biomass in comparison with the higher moisture regimes. The study also showed that P. dulce and A. nilotica seedlings were able to survive and maintain growth at low moisture regimes. The significant interaction effect in most parameters highlighted that both species and moisture regimes played an important role in germination and total biomass production, but this interaction had an insignificant effect on the survivability of all species. The result from this study highlights that species-specific responses to water stress could contribute to the species selection policies in drought-prone areas.

Allotetraploidization in Brachypodium May Have Led to the Dominance of One Parent's Metabolome in Germinating Seeds.

Seed germination is a complex process during which a mature seed resumes metabolic activity to prepare for seedling growth. In this study, we performed a comparative metabolomic analysis of the embryo and endosperm using the community standard lines of three annual Brachypodium species, i.e., B. distachyon (Bd) and B. stacei (Bs) and their natural allotetraploid B. hybridum (BdBs) that has wider ecological range than the other two species. We explored how far the metabolomic impact of allotetraploidization would be observable as over-lapping changes at 4, 12, and 24 h after imbibition (HAI) with water when germination was initiated. Metabolic changes during germination were more prominent in Brachypodium embryos than in the endosperm. The embryo and endosperm metabolomes of Bs and BdBs were similar, and those of Bd were distinctive. The Bs and BdBs embryos showed increased levels of sugars and the tricarboxylic acid cycle compared to Bd, which could have been indicative of better nutrient mobilization from the endosperm. Bs and BdBs also showed higher oxalate levels that could aid nutrient transfer through altered cellular events. In Brachypodium endosperm, the thick cell wall, in addition to starch, has been suggested to be a source of nutrients to the embryo. Metabolites indicative of sugar metabolism in the endosperm of all three species were not prominent, suggesting that mobilization mostly occurred prior to 4 HAI. Hydroxycinnamic and monolignol changes in Bs and BdBs were consistent with cell wall remodeling that arose following the release of nutrients to the respective embryos. Amino acid changes in both the embryo and endosperm were broadly consistent across the species. Taking our data together, the formation of BdBs may have maintained much of the Bs metabolome in both the embryo and endosperm during the early stages of germination. In the embryo, this conserved Bs metabolome appeared to include an elevated sugar metabolism that played a vital role in germination. If these observations are confirmed in the future with more Brachypodium accessions, it would substantiate the dominance of the Bs metabolome in BdBs allotetraploidization and the use of metabolomics to suggest important adaptive changes.

MicroRNAs expression dynamics reveal post-transcriptional mechanisms regulating seed development in Phaseolus vulgaris L.

The knowledge on post-transcriptional regulation mechanisms implicated in seed development (SD) is still limited, particularly in one of the most consumed grain legumes, Phaseolus vulgaris L. We explore for the first time the miRNA expression dynamics in P. vulgaris developing seeds. Seventy-two known and 39 new miRNAs were found expressed in P. vulgaris developing seeds. Most of the miRNAs identified were more abundant at 10 and 40 days after anthesis, suggesting that late embryogenesis/early filling and desiccation were SD stages in which miRNA action is more pronounced. Degradome analysis and target prediction identified targets for 77 expressed miRNAs. While several known miRNAs were predicted to target HD-ZIP, ARF, SPL, and NF-Y transcription factors families, most of the predicted targets for new miRNAs encode for functional proteins. MiRNAs-targets expression profiles evidenced that these miRNAs could tune distinct seed developmental stages. MiRNAs more accumulated at early SD stages were implicated in regulating the end of embryogenesis, postponing the seed maturation program, storage compound synthesis and allocation. MiRNAs more accumulated at late SD stages could be implicated in seed quiescence, desiccation tolerance, and longevity with still uncovered roles in germination. The miRNAs herein described represent novel P. vulgaris resources with potential application in future biotechnological approaches to modulate the expression of genes implicated in legume seed traits with impact in horticultural production systems.

Occurrence and Biosynthesis of Melatonin and Its Exogenous Effect on Plants

Melatonin is an endogenous indolamine found in many plants. It has been shown to generate a wide range of metabolic, physiological, and cellular responses, thus affecting growth and development, particularly under different environmental stresses. In the present review, we focus on its role in germination, growth and development, photosynthesis, senescence, and antioxidant activity in plants. Further, an effort has been made to discuss its occurrence, biosynthesis, and relationship with other phytohormones in plants. Moreover, melatonin-mediated signaling and its mechanisms of action under stress conditions in plants have been comprehensively discussed. Finally, its role under various abiotic stress conditions has also been discussed in this review.

Endophytic fungi associated with roots of epiphytic orchids in two Andean forests in Southern Ecuador and their role in germination

Orchids are known to establish complex relationships with endophytic fungi throughout their life cycle, and particularly during germination of their reserves-deprived seeds. Characterizing generalist or specialist interactions between orchids and associated fungi is key to supporting orchid conservation efforts. Here, endophytic fungi associated with roots of epiphytic orchids were studied in two montane Andean forests in Southern Ecuador. Orchid root samples were collected from ten ~500 m2 plots distributed between two neighboring forests. Endophytic fungi associated with these roots were then isolated, cultured, and identified by sequencing of rDNA markers. In total, 52 pure isolates were recovered from the roots of 10 orchid species. These isolates were classified into nine taxonomic groups except for one isolate that remained unclassified. Most fungal isolates were found in roots of up to two different orchid species; however, Coprinellus radians was found in the roots of all sampled orchids. The potential of C. radians to promote germination of orchid seeds was tested in a separate assay using seeds from two orchid species different than those found in the experimental forest plots. Of the two C. radians isolates tested, one improved germination in the two orchids evaluated to a level about half of that observed in seeds germinated in nutrient-rich medium (Phytamax) and above the null germination observed in plates without the fungus. Together, these results revealed a generalist relationship between C. radians and all the studied epiphytic orchids and the potential role of this fungus as a promoter of orchid seed germination.&#x0D; Key Words: Agaricales, Epidendrum, symbiotic germination

Effects of Foliar Application of Indol Butyric Acid(IBA), Gibberellic Acid (GA3) and Zinc (Zn) on Yield and Quality of Tomato

The yield and quality of tomato largely depend on soil and climatic conditions and also on variety. Plant growth regulators function as chemical messengers for intercellular communication. In tomato, different growth regulators and zinc play a pivotal role in germination, root development, branching, flower initiation, fruiting, lycopene development, synchronization and early maturation, parthenocarpic fruit development, ripening, TSS, acidity, seed production etcetera. The field experiment was conducted at Sher-e-Bangla Agricultural University Farm, Dhaka, Bangladesh during October 2017 to April, 2018 to evaluate the effect of foliar application of IBA, GA3 and Zn on yield and quality of tomato [1]. Single variety BARI Tomato-14 and foliar application of IBA 5 ml/L, GA3 5 ml/l and Zn were used to conduct this experiment. The experiment was laid out in Randomized Complete Block Design (RCBD) having three factors and replicated three times. Data were collected on chlorophyll content of leaf, number of flowers cluster per pant, number of fruit per plant, yield per hectare, Vitamin-C content and TSS% content. A statistically significant variation was recorded in terms of the above characters about yield and quality of tomato. All the characters shows maximum result in treatment of T7 (IBA+GA3+Zn).The treatment combination of T7 (IBA 5 ml / L, GA3 5 ml / L and Zn 1 kg/ha) and T4 (IBA 5 ml / L, GA3 5 ml / L) gave the maximum yield (100.00 t/ha) and the minimum yield (39.75 t/ha) was found from the treatment combination on T0 (control). The highest Vitamin C content (113.10 mg/100 gm) and highest TSS (%) (7.000) were found from T7 (IBA 5 ml / L, GA3 5 ml / L and Zn 1 kg/ha) than that of control under present experiment. So, it can be concluded that treatment combination of T7 (IBA 5 ml / L, GA3 5 ml / L and Zn 1 kg/ha) is the best for yield and quality of tomato and can be tested further under different field conditions.

Climate drives differences in the germination niche of a globally distributed invasive grass

Abstract Aims Exotic invasive species are often exposed to strong selection pressures in their new ranges that can often lead to substantial intraspecific variation. Population differentiation in the timing of life history events in response to climate gradients is thought to be an important mechanism facilitating the range expansion of many invasive species. For seed producing plants, the timing of seed germination determines the first environmental conditions experienced by newly emerged germinates, and can have important implications for the successful colonization, establishment and spread of invasive plants—though the role of germination in the success of invasive plants remains poorly understood. Methods We assessed the variation in seed germination dynamics among 10 populations of the invasive plant Johnsongrass (Sorghum halepense) across its North American distribution, capturing both a temperature and precipitation gradient, and whether that variation is associated with home climate. Seeds were exposed to a wide range of temperatures (11–48°C) and two water availability treatments. Important Findings We found that Johnsongrass seeds germinated across a wide range of temperatures, but there was substantial variation among populations in the proportion of seeds that germinated in response to both temperature and water availability. Evidence indicates that as Johnsongrass expanded its range from warmer climates into cooler climates, there was a concurrent shift in the germination temperature niche to cooler temperatures. Our results suggest that the germination of Johnsongrass seeds has adapted to home climate allowing this invader to maximize germination throughout its range, and that this may be an important contributing factor to its invasion into new environments.

English

Upland rice is an ecotype adapted to dry culture system. In this study, changes in germination characteristics and transcriptome of germinating seeds exposed to 15% polyethylene glycol (PEG) were investigated using upland rice genotype IRAT109. PEG reduced germination potential and inhibited growth of seed radicle and plumule. Under PEG stress, gibberellic acid (GA) content decreased and abscisic acid (ABA) content increased. Transcriptome analysis revealed that 1270 genes were differentially expressed between stressed and non-stressed seeds. Approximately, 56.46% of differentially expressed genes (DEGs) were up-regulated and 43.54% of DEGs were down-regulated under PEG stress. Gene Ontology (GO) analysis categorized 1149 DEGs into 52 functional groups and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis assigned 123 DEGs to 54 pathways. A large number of DEGs related to GA signal transduction, ABA biosynthesis and signal transduction, and defense response pathways were upregulated. Results indicated that genes related with GA signal transduction and ABA play important role in germination under drought stress. Findings of this study are highly useful to understand the molecular mechanism of germination and provide candidate genes for molecular breeding in dry direct-seeded rice. © 2019 Friends Science Publishers

A CCR4 Association Factor 1, OsCAF1B, Participates in the αAmy3 mRNA Poly(A) Tail Shortening and Plays a Role in Germination and Seedling Growth.

Poly(A) tail (PAT) shortening, also termed deadenylation, is the rate-limiting step of mRNA degradation in eukaryotic cells. The carbon catabolite repressor 4-associated factor 1s (CAF1s) were shown to be one of the major enzymes for catalyzing mRNA deadenylation in yeast and mammalian cells. However, the functions of CAF1 proteins in plants are poorly understood. Herein, a sugar-upregulated CAF1 gene, OsCAF1B, is investigated in rice. Using gain-of-function and dominant-negative mutation analysis, we show that overexpression of OsCAF1B resulted in an accelerated α-amylase gene (αAmy3) mRNA degradation phenomenon, while ectopic expression of a form of OsCAF1B that had lost its deadenylase activity resulted in a delayed αAmy3 mRNA degradation phenomenon in transgenic rice cells. The change in αAmy3 mRNA degradation in transgenic rice is associated with the altered lengths of the αAmy3 mRNA PAT, indicating that OsCAF1B acts as a negative regulator of αAmy3 mRNA stability in rice. Additionally, we found that overexpression of OsCAF1B retards seed germination and seedling growth. These findings indicate that OsCAF1B participates in sugar-induced αAmy3 mRNA degradation and deadenylation and acts a negative factor for germination and seedling development.

Identification and Characterization of Thioredoxin H-Type Gene Family in Triticum turgidum ssp. durum in Response to Natural and Environmental Factor-Induced Oxidative Stress

Key message: Thioredoxin h-type isoforms are tissue-specific, differentially expressed in germinating seeds and under salinity stress and highly regulated by H2O2, suggestive of a role in germination and salinity adaptation in durum wheat through redox regulation. Thioredoxins (Trxs) are protein-disulfide reductases that perform multiple functions related to cellular redox homoeostasis. The most complex cluster in the family of plant Trxs is formed by h-type Trxs since their identity and functions are still largely unknown. Here, h-type Trxs from durum wheat (Triticum turgidum ssp. durum) (stated TdTrxh1, TdTrxh2, TdTrxh3, and TdTrxh9) were identified and characterized in response to oxidative stress either generated naturally (germination) or induced by salinity and hydrogen peroxide (H2O2). In silico expression analysis, based on RNA-seq data, revealed a tissue-specific expression for TdTrxh genes. Real-time q-PCR analysis showed differential expression patterns for TdTrxh genes between and within seed tissues during germination and a marked induction of TdTrxh2 and h3 ones. Moreover, the four TdTrxh isoforms were found to be differentially modulated by salinity in two durum wheat varieties contrasting in their salinity tolerance. Particularly, an upregulation of the four TdTrxh genes (mainly TdTrxh2 and h3) was noted in root tissues of the tolerant variety, while they were markedly downregulated in those of the sensitive one. This upregulation of TdTrxh genes in the tolerant variety coincided with an accumulation of H2O2 in root tissues. When exogenously applied, H2O2 increased mRNA transcripts of all TdTrxh genes in both varieties regardless of their salinity-tolerance degree. Collectively, our data suggest a non-redundant function for h-type Trxs besides to be potentially involved in salinity tolerance in durum wheat through redox regulation.

Cytological and histochemical evaluations reveal roles of the cotyledonary petiole in the germination and seedling development of Mauritia flexuosa (Arecaceae)

The cotyledonary petiole (CP) completely envelops the embryo axis during embryogenesis in Arecaceae. There is little information available, however, on the roles of that structure in seed germination and initial seedling development-crucial plant life cycle phases. The study therefore sought to evaluate the roles of CP in the germination and post-seminal development of the recalcitrant seeds of Mauritia flexuosa, an ecologically and economically important neotropical palm. The CP and the embryo/vegetative axis were evaluated during germination and initial seedling development using standard morphological, anatomical, histochemical, and ultrastructural methodologies. Evaluations of dormant seeds incubated for 60days were also performed. The CP (a) promotes seedling protrusion in the germination, extending the embryo axis outside the seed; (b) protects the vegetative axis through the development of coating rich in phenolic compounds and lignin; (c) participates in reserve translocation, with the conversion of its own proteinaceous/mucilaginous reserves into transitional starch, as well as acting in the transport of endospermic reserves; (d) favors aeration, with the formation of pathways among stomata, substomatal chambers, and intercellular spaces; (e) controls seedling morphogenesis by modulating the curvature of the vegetative axis; and (f) contributes to overcoming seed bank dormancy through cytological alterations (protein synthesis and mitochondrial proliferation). The cotyledonary petiole of palms is a unique and multifunctional structure among angiosperms, with crucial roles in germination and seedling establishment.

Seed germination and dormancy: The classic story, new puzzles, and evolution.

This review highlights recent progresses in seed germination and dormancy research. Research on the weakening of the endosperm during germination, which is almost a classic theme in seed biology, was resumed by α-xylosidase studies. Strong genetic evidence was presented to suggest that the quality control of xyloglucan biosynthesis in the endosperm (and the embryo) plays a critical role in germination. Further analyses on the endosperm and the adjacent layers have suggested that the cutin coat in the endosperm-testa interphase negatively affects germination while the endosperm-embryo interphase produces a sheath that facilitates germination. These progresses significantly advanced our understanding of seed germination mechanisms. A breakthrough in dormancy research, on the other hand, revealed the unique abscisic acid signaling pathway that is regulated by DELAY OF GERMINATION1 (DOG1). The detailed analysis of DOG1 expression uncovered the intriguing story of reciprocal regulation of the sense-antisense pair, which generated new questions. Recent studies also suggested that the DOG1 function is not limited to dormancy but extended through general seed maturation, which provokes questions about the evolution of DOG1 family proteins. Seed biology is becoming more exciting with the classic stories being revitalized and new puzzles emerging from the frontier.

New Insight on Water Status in Germinating Brassica napus Seeds in Relation to Priming-Improved Germination.

Seed priming is a pre-sowing method successfully used to improve seed germination. Since water plays a crucial role in germination, the aim of this study was to investigate the relationship between better germination performances of osmoprimed Brassica napus seeds and seed water status during germination. To achieve this goal, a combination of different kinds of approaches was used, including nuclear magnetic resonance (NMR) spectroscopy, TEM, and SEM as well as semi-quantitative PCR (semi-qPCR). The results of this study showed that osmopriming enhanced the kinetics of water uptake and the total amount of absorbed water during both the early imbibition stage and in the later phases of seed germination. The spin–spin relaxation time (T2) measurement suggests that osmopriming causes faster water penetration into the seed and more efficient tissue hydration. Moreover, factors potentially affecting water relations in germinating primed seeds were also identified. It was shown that osmopriming (i) changes the microstructural features of the seed coat, e.g., leads to the formation of microcracks, (ii) alters the internal structure of the seed by the induction of additional void spaces in the seed, (iii) increases cotyledons cells vacuolization, and (iv) modifies the expression pattern of aquaporin genes.

Characterization of regulatory elements in OsRGLP2 gene promoter from different rice accessions through sequencing and in silico evaluation

Germins and germin-like proteins from cupin superfamily contribute resistance to heat denaturation, chemical degradation and against plant pathogens, further functions in plant growth and development. In this study, from three different Oryza sativa accessions KS-282 and Pak 7178 and Pak 7865, OsRGLP2 gene promoter region was amplified, sequenced and analyzed. Sequencing data was evaluated via different computational tools. The regulatory elements were predicted by Consite tool and mapping was done. Many transcription factors binding sites were discovered in OsRGLP2 gene promoter; among these factors, HFH-1 having a significant role in germination was picked for further investigation. To study the interaction between HFH-1 and corresponding regulatory factors, HADDOCK Webserver was used. Graphical models for the interactions of HFH-1 and related regulatory elements were studied by graphic molecular system PyMOL. Mapping of cis-acting regulatory elements in OsRGLP2 gene promoter from three rice accessions showed differences in their position and copy number. Important regulatory elements found in OsRGLP2 promoter region were TATA, CAAT Box, ARR1, GATA, AGAAA, CAAT and DNA-binding One Zinc Finger (Dof) factors, few of them contribute to the regulation of plant defensive system, light responses, developmental and growth activities. Furthermore, during DNA interaction studies, it was found that HFH-1 transcription factor participates in hydrogen bonds formation with thymine and adenine bases.

Utilisation of stored lipids during germination in dimorphic seeds of euhalophyte Suaeda salsa.

Utilisation of stored lipids plays an important role in germination of oil seeds. In the present study, key enzyme activity (lipase, isocitrate lyase and malate synthase) in lipid utilisation was determined during germination in dimorphic seeds of euhalophyte Suaeda salsa (L.) Pall. The results revealed that the percentage of germination were highest in intertidal brown seeds, followed by inland brown seeds and then inland black seeds moistened with 0 and 300mM NaCl during early seed germination. The same trend was found in the activity of three enzymes and soluble sugar content when seeds were moistened with 0 and 300mM NaCl for 3h. Salinity reduced the activity of three enzymes in inland brown and black seeds in the initial 3h, except that salinity had no adverse effect on isocitrate lyase activity of brown seeds. Salinity had no adverse effect on three enzymes in inland brown and black seeds in the initial 30h, except that it decreased malate synthase activity of black seeds. Salinity had no effect on three enzymes in intertidal brown seeds in the initial 3h and 30h. In conclusion, high activity of these enzymes in brown seeds may play an important role in utilisation of stored lipids during their rapid seed germination.

Involvement of nitric oxide in enhanced germination and seedling growth of magnetoprimed maize seeds

ABSTRACTNitric oxide (NO) is a highly reactive gaseous free radical, which in plants was found to stimulate seed germination and ending of dormancy. Experiments were conducted to study the effect of NO inhibitors sodium tungstate (ST) and Nω-nitro-L-arginine methyl ester hydrochloride (L-NAME), NADPH oxidase inhibitor diphenyleneiodonium (DPI) and NO donor sodium nitroprusside (SNP) on untreated and magnetoprimed maize (Zea mays var: GSF-2) seeds. Treatment of maize seeds with these inhibitors inhibited germination related parameters like seedling length, fresh weight, dry weight and vigour indices and α-amylase activity of maize seeds under laboratory conditions, whereas NO donor (SNP) promoted all these parameters. Among 3 different inhibitors used ST was most effective and showed an inhibition in seedling length of 67% and 71% at 1 mM concentration for untreated (UT) and magnetically treated (MT) seeds respectively. Data presented here indicate the involvement of nitric oxide in enhanced germination and seedling growth of magnetoprimed maize seeds. ROS are continuosly produced by the cells of germinating seeds and play a positive role in germination of magnetoprimed maize seeds. ROS inhibitor (DPI) inhibited seedling length by 34% and 40% for control and MT seeds respectively. α-amylase activity was also inhibited by all the 3 inhibitors used. It is concluded that NO inhibitors and ROS inhibitor inhibited magnetic field induced promotion of seedling parameters and α- amylase activity of maize seeds.

Canonical germinant receptor is dispensable for spore germination in Clostridium botulinum group II strain NCTC 11219

Clostridium botulinum is an anaerobic sporeforming bacterium that is notorious for producing a potent neurotoxin. Spores of C. botulinum can survive mild food processing treatments and subsequently germinate, multiply, produce toxin and cause botulism. Control of spore germination and outgrowth is therefore essential for the safety of mildly processed foods. However, little is known about the process of spore germination in group II C. botulinum (gIICb), which are a major concern in chilled foods because they are psychrotrophic. The classical model of spore germination states that germination is triggered by the binding of a germinant molecule to a cognate germinant receptor. Remarkably, unlike many other sporeformers, gIICb has only one predicted canonical germinant receptor although it responds to multiple germinants. Therefore, we deleted the gerBAC locus that encodes this germinant receptor to determine its role in germination. Surprisingly, the deletion did not affect germination by any of the nutrient germinants, nor by the non-nutrient dodecylamine. We conclude that one or more other, so far unidentified, germinant receptors must be responsible for nutrient induced germination in gIICb. Furthermore, the gerBAC locus was strongly conserved with intact open reading frames in 159 gIICb genomes, suggesting that it has nevertheless an important function.

Dominant Groups of Potentially Active Bacteria Shared by Barley Seeds become Less Abundant in Root Associated Microbiome.

Endophytes are microorganisms colonizing plant internal tissues. They are ubiquitously associated with plants and play an important role in plant growth and health. In this work, we grew five modern cultivars of barley in axenic systems using sterile sand mixture as well as in greenhouse with natural soil. We characterized the potentially active microbial communities associated with seeds and roots using rRNA based amplicon sequencing. The seeds of the different cultivars share a great part of their microbiome, as we observed a predominance of a few bacterial OTUs assigned to Phyllobacterium, Paenibacillus, and Trabusiella. Seed endophytes, particularly members of the Enterobacteriacea and Paenibacillaceae, were important members of root endophytes in axenic systems, where there were no external microbes. However, when plants were grown in soil, seed endophytes became less abundant in root associated microbiome. We observed a clear enrichment of Actinobacteriacea and Rhizobiaceae, indicating a strong influence of the soil bacterial communities on the composition of the root microbiome. Two OTUs assigned to Phyllobacteriaceae were found in all seeds and root samples growing in soil, indicating a relationship between seed-borne and root associated microbiome in barley. Even though the role of endophytic bacteria remains to be clarified, it is known that many members of the genera detected in our study produce phytohormones, shape seedling exudate profile and may play an important role in germination and establishment of the seedlings.

A Clostridium difficile alanine racemase affects spore germination and accommodates serine as a substrate

Clostridium difficile has become one of the most common bacterial pathogens in hospital-acquired infections in the United States. Although C. difficile is strictly anaerobic, it survives in aerobic environments and transmits between hosts via spores. C. difficile spore germination is triggered in response to certain bile acids and glycine. Although glycine is the most effective co-germinant, other amino acids can substitute with varying efficiencies. Of these, l-alanine is an effective co-germinant and is also a germinant for most bacterial spores. Many endospore-forming bacteria embed alanine racemases into their spore coats, and these enzymes are thought to convert the l-alanine germinant into d-alanine, a spore germination inhibitor. Although the C. difficile Alr2 racemase is the sixth most highly expressed gene during C. difficile spore formation, a previous study reported that Alr2 has little to no role in germination of C. difficile spores in rich medium. Here, we hypothesized that Alr2 could affect C. difficile l-alanine-induced spore germination in a defined medium. We found that alr2 mutant spores more readily germinate in response to l-alanine as a co-germinant. Surprisingly, d-alanine also functioned as a co-germinant. Moreover, we found that Alr2 could interconvert l- and d-serine and that Alr2 bound to l- and d-serine with ∼2-fold weaker affinity to that of l- and d-alanine. Finally, we demonstrate that l- and d-serine are also co-germinants for C. difficile spores. These results suggest that C. difficile spores can respond to a diverse set of amino acid co-germinants and reveal that Alr2 can accommodate serine as a substrate.

Fungal endophytes from seeds of invasive, non-native Phragmites australis and their potential role in germination and seedling growth

We characterized fungal endophytes of seeds of invasive, non-native Phragmites from three sites in the Great Lakes region to determine if fungal symbiosis could contribute to invasiveness through their effects on seed germination and seedling growth. Field-collected seeds were surface sterilized and plated on agar to culture endophytes for ITS sequencing. Prevalence of specific endophytes from germinated and non-germinated seeds, and from seedlings, was compared. One-third of 740 seeds yielded endophyte isolates. Fifteen taxa were identified with Alternaria sp. representing 54% of all isolates followed by Phoma sp. (21%) and Penicillium corylophilum (12%). Overall germination of seeds producing an isolate (36%) was significantly higher than seeds not producing an isolate (20%). Penicillium in particular was strongly associated with increased germination of seeds from one site. Sixty-three isolates and 11 taxa were also obtained from 30 seedlings where Phoma, Penicillium and Alternaria respectively were most prevalent. There was a significant effect of isolating an endophyte from the seed on seedling growth. These results suggest that many endophyte taxa are transmitted in seeds and can increase seed germination and seedling growth of invasive Phragmites. The role of fungal endophytes in host establishment, growth and invasiveness in nature requires further research.