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Antimicrobial resistance is a growing concern in public health and current research shows an important role for bacterial biofilms in recurrent or chronic infections. New strategies, therefore, are necessary to overcome antimicrobial resistance, through the development of new therapies that could alter or inhibit biofilm formation. In this sense, antibiofilm natural products are very promising. In this work, a bioprospection of antimicrobial and antibiofilm extracts from Uruguayan soil bacteria and insect gut bacteria was carried out. Extracts from extracellular broths were tested for their ability to inhibit planktonic cell growth and biofilm formation. Genomic analysis of Bacillus cereus ILBB55 was carried out. All extracts were able to inhibit the growth of, at least, one microorganism and several extracts showed MICs lower than 500 µg mL−1 against microorganisms of clinical relevance (Staphylococcus aureus, Pseudomonas aeruginosa, and Enterobacter cloacae). Among the extracts evaluated for biofilm inhibition only ILBB55, from B. cereus, was able to inhibit, S. aureus (99%) and P. aeruginosa (62%) biofilms. Genomic analysis of this strain showed gene clusters similar to other clusters that code for known antimicrobial compounds. Our study revealed that extracts from soil bacteria and insect gut bacteria, especially from B. cereus ILBB55, could be potential candidates for drug discovery to treat infectious diseases and inhibit S. aureus and P. aeruginosa biofilms.

Claviceps species affecting Paspalum spp. are a serious problem, as they infect forage grasses such as Paspalum dilatatum and P. plicatulum, producing the ergot disease. The ascomycete C. paspali is known to be the pathogen responsible for this disease in both grasses. This fungus produces alkaloids, including ergot alkaloids and indole-diterpenes, that have potent neurotropic activities in mammals. A total of 32 isolates from Uruguay were obtained from infected P. dilatatum and P. plicatulum. Isolates were phylogenetically identified using partial sequences of the genes coding for the second largest subunit of RNA polymerase subunit II (RPB2), translation elongation factor 1-α (TEF1), β-tubulin (TUB2), and the nuc rDNA 28S subunit (28S). Isolates were also genotyped by randomly amplified polymorphic DNA (RAPD) and presence of genes within the ergot alkaloid (EAS) and indole-diterpene (IDT) biosynthetic gene clusters. This study represents the first genetic characterization of several isolates of C. paspali. The results from this study provide insight into the genetic and genotypic diversity of Claviceps paspali present in P. dilatatum and suggest that isolates from P. plicatulum could be considered an ecological subspecies or specialized variant of C. paspali. Some of these isolates show hypothetical alkaloid genotypes never reported before.

SummaryInsect Epicuticle hydrocarbons (CHC) are known to be important determinants in the susceptibility degree of insects to fungal entomopathogens. Five Beauveria bassiana isolates were phenotypically analyzed regarding their response to CHC nutrition and their pathogenicity and virulence towards high fungal-susceptible Thaumastocoris peregrinus and low fungal-susceptible Piezodorus guildinii, which are important hemipteran pests in eucalyptus and soybean plantations, respectively. Two of these isolates, resulting the most (ILBB308) and the least (ILBB299) virulent to P. guildinii, were also evaluated at gene expression level after growth on n-pentadecane. B. bassiana most virulent isolate ILBB308 showed the lowest growth on most evaluated CHC media. However, this isolate distinctively induced most of the analyzed genes involved in CHC assimilation, cuticle degradation and stress tolerance. Virulence towards low susceptibility P. guildinii was enhanced in both hypervirulent ILB308 and hypovirulent ILBB299 isolates after growth on n-pentadecane as the sole carbon source, whereas virulence enhancement towards high susceptibility T. peregrinus was not observed in alkane-grown fungi. Virulence enhancement towards P. guildinii could be mostly explained by a priming effect produced by CHC on the induction of some genes related to hydrocarbon assimilation in ILB 205 and ILB 308, such as hydrophobin (Bbhyd2) and cytochrome P450 genes (BbCyp52g11 and BbCyp52×1), and partially by the induction of genes related to cuticle degradation (Bbchit and Bbcdep1) and stress tolerance (Bbsod1) observed only in ILB308.

BACKGROUND The redbanded stink bug Piezodorus guildinii (Heteroptera: Pentatomidae) is one of the most important species affecting soybean crops in southern South America. Capillary gas chromatography coupled to mass spectrometry was used to characterize the epicuticular hydrocarbon profiles of field-collected insects, and to identify differences in their composition between fifth-instar nymphs and adults, males and females, and between bugs collected in insecticide-treated and insecticide-free soybean crops. RESULTS Straight chain saturated n-C27 and n-C29, and monomethyl and dimethyl chains of C31 and C33 were the most abundant compounds. A group of volatile hydrocarbons with n-C13 and n-C15 as the predominant compounds were also detected. The hydrocarbon pattern was different between nymphs and adults, either males or females. Heneicosene was almost exclusively detected in adult males and was the most important component to differentiate between both sexes, followed by tricosadiene. The total hydrocarbon amount was significantly higher in nymphs, males and females collected in insecticide-treated fields compared with insects obtained from untreated fields. CONCLUSION Differences were found in the epicuticular hydrocarbon pattern among nymphs and adults, as well as sexual dimorphism in adult stink bugs. Interestingly, an alteration was also found in the hydrocarbon profile of insects collected in insecticide-treated soybean crops and its relevance is discussed within a pest management context.

Thaumastocoris peregrinus Carpintero & Dellape (Heteroptera: Thaumastocoridae) is a sap‐sucking insect that feeds on leaves of Eucalyptus. In Uruguay, it was detected in 2008 causing significant economic losses in Eucalyptus plantations. At present, there is no efficient control for this pest; thus, the use of biological control agents seems to be an environmentally friendly alternative to reduce the damage caused by this insect. The aims of this study were to isolate and identify the species of entomopathogenic fungi that naturally infect T. peregrinus in Uruguay and to characterize and select the most virulent isolates towards this pest. Individuals of T. peregrinus were collected in eight Eucalyptus plantations infested by the pest. The entomopathogenic fungi were isolated and identified by observation of their micromorphological characteristics, and their identity was confirmed by molecular methods. The pathogenicity and virulence against T. peregrinus of the isolated entomopathogenic fungi were evaluated. Isolates causing the highest insect mortality were selected to evaluate the effect of temperature and water activity on conidial viability. Entomopathogenic fungi were found in very low number with the prevalence of less than 3%. Isolates corresponded to Beauveria, Fusarium, Isaria, Lecanicillium, Paecilomyces, Pochonia, Purpureocillium and Simplicillium genera. Two species, B. pseudobassiana and L. muscarium, were first recorded in Uruguay. Among all the tested species, an isolate of B. bassiana (FI 2403) showed the highest virulence followed by an isolate of B. pseudobassiana. The isolate of B. bassiana presented the highest percentage of spore germination at the three temperatures and the highest viability at low water activities. Isolate FI 2403 was selected as a promissory candidate for the development of a commercial formulation against T. peregrinus.

The bronze bug Thaumastocoris peregrinus is an invasive pest, affecting Eucalyptus plantations worldwide. Although its natural enemy Cleruchoides noackae has been tested for the biological control of this pest, other strategies like the use of native entomopathogenic fungi are needed. For this, native virulent fungal isolates should be selected, massively multiplied in an efficient way, and prepared to obtain a stable product. Isolates of native Beauveria bassiana obtained from T. peregrinus and from different collections were screened for their virulence towards this insect and for their amenability to be massively produced in a low-input liquid submerged fermentation and prepared as a dry powder. Three out of six virulent strains were suitable for their massive production in a 2% corn flour suspension, achieving 109 submerged propagules/g of dehydrated preparation. The LC50 achieved by the dry submerged propagules did not differ from the LC50 of fresh aerial conidia. The proposed dual selection ...

Phosphorus deficiency can be a major limitation to legume growth when plant nitrogen nutrition depends on symbiotic nitrogen fixation. One possible approach to overcome this constraint is the selection of plant and rhizobial genotypes capable of metabolizing complex forms of phosphorus in the nodules. The aim of this research was to study the rhizobial phytase transcript abundance in nodules of two soybean cultivars (Glycine max (L.) Merr.) grown under two different phosphorus conditions in hydroaeroponic conditions. An in situ RT-PCR of a rhizobial phytase was performed in microtome sections of soybean nodules of two cultivars growing under phosphorus sufficiency vs. phosphorus deficiency. The results showed that the plant cultivar may influence the level of transcript abundance of the bacterial phytase and in consequence affect the phosphorus use efficiency of nitrogen-dependent Bradyrhizobium spp.-soybean symbioses. Thus, the selection of a good combination of plant and rhizobial genotypes should be a priority when breeding for phosphorus deficiency is performed.

Serratia marcescens can be a plant growth promoting bacteria (PGPB) and an opportunistic human and plant pathogen. We have identified and characterized strains of related species of Serratia and evaluated their biological control of damping-off of tomato seeds caused by Pythium cryptoirregulare. Serratia ureilytica, S. bockelmannii and S. nevei were identified by phylogenetic analysis of partial gyrB gene sequence and average nucleotide identity (ANI). Tomato seeds inoculated with S. ureilytica ILBB 145 showed higher germination percentage and reduced damping-off in greenhouse experiment resembling a commercial operation, and volatiles produced by this strain caused the nearly complete inhibition in vitro of P. cryptoirregulare. Analysis of volatile organic compounds (VOCs) showed that ILBB 145 produced dimethyl disulfide (DMDS), which can partially account for this inhibition. Serratia bockelmannii ILBB 162 performance against damping-off was intermediate and the inhibition of P. cryptoirregulare in vitro was lower and explained by volatile and diffusible metabolites. Both strains augmented DMDS production in the presence of P. cryptoirregulare, suggesting this compound may play a role in the context of interspecific competition. Serratia nevei ILBB 219 showed the lowest inhibition of P. cryptoirregulare in vitro, no DMDS production, and no biocontrol in planta. Draft genomes of the three strains were annotated and individual genes and biosynthesis gene clusters were identified in relation with the observed phenotypes. We report S. ureilytica - a low risk species- with activity as a biological control agent and DMDS produced by this bacterial species putatively involved in seed and seedling protection against P. cryptoirregulare.

Soil microbial communities play critical roles in maintaining natural ecosystems such as the Campos biome grasslands of southern South America. These grasslands are characterized by a high diversity of soils, low available phosphorus (P) and limited water holding capacity. This work aimed to describe prokaryotic communities associated with different soil types and to examine the relationship among these soil communities, the parent material and the soil nutrient status. Five Uruguayan soils with different parent material and nutrient status, under natural grasslands, were compared. The structure and diversity of prokaryotic communities were characterized by sequencing 16S rRNA gene amplicons. Proteobacteria, Actinobacteria, Firmicutes,Verrucomicrobia, Acidobacteria, Planctomycetes and Chloroflexi were the predominant phyla. Ordination based on several distance measures was able to discriminate clearly between communities associated with different soil types. Edge-PCA phylogeny-sensitive ordination and differential relative abundance analyses identified Archaea and the bacterial phyla Firmicutes, Acidobacteria, Actinobacteria and Verrucomicrobia as those with significant differences among soil types. Canonical analysis of principal coordinates identified porosity, clay content, available P, soil organic carbon and water holding capacity as the main variables contributing to determine the characteristic prokaryotic communities of each soil type.

Insect Epicuticle hydrocarbons (CHC) are known to be important determinants in the susceptibility degree of insects to fungal entomopathogens. Five Beauveria bassiana (Balsamo) Vuillemin (Hypocreales; Clavicipitaceae) strains were phenotypically analyzed regarding their response to CHC nutrition and their pathogenicity and virulence towards high fungal-susceptible Thaumastocoris peregrinus (Carpintero and Dellape) (Heteroptera: Thaumastocoridae) and low fungal-susceptible Piezodorus guildinii (Westwood) (Hemiptera: Pentatomidae), which are important hemipteran pests in eucalyptus and soybean plantations, respectively. Two of these strains, which were the most (ILB308) and the least (ILB299) virulent to P. guildinii, were also evaluated at gene expression level after growth on n-pentadecane, a P. guildinii epicuticular hydrocarbon. Beauveria bassiana hypervirulent strain ILB308 showed the lowest growth on most evaluated CHC media. However, this strain distinctively induced most of the analyzed genes involved in CHC assimilation, cuticle degradation and stress tolerance. Virulence towards low susceptibility P. guildinii was enhanced in both hypervirulent ILB308 and hypovirulent ILB299 strains after growth on n-pentadecane as the sole carbon source, whereas virulence enhancement towards high susceptibility T. peregrinus was only observed in the hypervirulent strain. Virulence enhancement towards P. guildinii could be mostly explained by a priming effect produced by CHC on the induction of some genes related to hydrocarbon assimilation in ILB299 and ILB308, such as cytochrome P450 genes (BbCyp52g11 and BbCyp52x1), together with adhesion and stress tolerance genes, such as hydrophobin (Bbhyd2) and catalase (Bbcatc) and glutathione peroxidase (Bbgpx), respectively.

Diseases remain a yield-limiting factor for crops despite the availability of control measures for many pathogens. Indigenous soil microorganisms can suppress some plant pathogens, yet there is little systematic information on the effects of cropping systems on disease-suppressive populations in soil. Streptomyces have been associated with suppression of plant diseases in several naturally occurring disease-suppressive soils. Pathogen-suppressive activity of Streptomyces communities is correlated with higher bacterial densities and with inhibitory phenotypes, driven by competition among indigenous soil bacteria. We sought to characterize relationships between cropping practices and pathogen suppression among soil Streptomyces. We evaluated bacterial and Streptomyces densities and inhibitory activities in soils from a long-term crop rotation experiment. Signaling interactions that altered inhibitory phenotypes among sympatric populations were also evaluated for a subset of samples. Soils from longer rotations, which had a higher number of plant species over time, had larger bacterial and Streptomyces densities, and more inhibitors than soils from shorter rotations. In addition, signaling occurred more frequently among isolates from higher-density communities. Our work shows that bacterial density, pathogen suppression and signaling are interrelated and are affected by crop rotation, suggesting the potential for management to optimize suppressive populations.

Teratosphaeria nubilosa severely affects young plantations of Eucalyptus globulus and Eucalyptus maidenii in Uruguay. This work analyzes the effect of the damage caused by T. nubilosa on tree growth and survival of E. globulus and E. maidenii, based on data from a trial naturally infected by this pathogen. The effect of defoliation and proportion of adult foliage on tree growth and mortality was evaluated at 50 months of age. Defoliation in E. globulus was higher than in E. maidenii, reaching average values of 52.5 and 28.8 %, respectively, at 21 months of age. By contrast, E. globulus showed higher precocity of vegetative phase change than E. maidenii. Although the damage threshold for diameter growth was 40 % on E. globulus and 20 % on E. maidenii, the detrimental effect on growth was similar in both species, where the most defoliated trees suffered a reduction in diameter of 49.4 and 47.5 %, respectively, at 50 months of age. Mortality increased significantly when defoliation exceeded 30 % in E. globulus and 40 % in E. maidenii. The results showed that E. maidenii has lower defoliation, higher growth and lower mortality than E. globulus after epidemics of T. nubilosa. However, the lower defoliation and higher proportion of juvenile foliage showed by E. maidenii represent a greater source of inoculum, suggesting that a large-scale replacement of E. globulus by E. maidenii in areas where T. nubilosa is a problem could be risky from an epidemiological point of view.

Pythium spp. were isolated from symptomatic plants and agricultural soils in Uruguay. Species were identified and the pathogenicity of representative isolates was assessed in vitro on lotus, alfalfa and soybean seeds. Pythium sylvaticum and P. paroecandrum were the least virulent on lotus and alfalfa, while P. irregulare, P. ultimum and P. cryptoirregulare were the most virulent. Soybean seeds were equally affected by isolates of all tested Pythium species.

Over the last 5 years foliar diseases had caused devastating damages in young plantations of Eucalyptus globulus in Uruguay. The sustainability of this species, the most important in the country with more than 270,000 ha of commercial plantations, depends on the rapid development of resistant genetic stock. The genetic variation in resistance of juvenile foliage to diseases and in the onset to adult foliage were examined in a field trial of E. globulus naturally infected by Mycosphaerella leaf disease (Mycosphaerella spp. and Teratosphaeria spp.) and Eucalyptus rust (Puccinia psidii). The genetic material included 226 open pollinated seed lots from 16 provenances of Australia, Chile and Uruguay. Disease severity (% of leaf spots and % defoliation) was assessed 8 months after planting and precocity of phase change (% of adult foliage) at 20 months. Tree growth and survival were evaluated every 2 years up to the ninth year. Differences in disease severity and in precocity of phase change were significant among countries of origin and provenances. Within provenances, the narrow-sense individual tree heritabilities for leaf spot severity (0.37), defoliation (0.41) and proportion of adult foliage (0.40) were higher than those obtained for tree growth (0.19 to 0.33) and similar to those obtained for survival (0.33 to 0.45). Genetic correlations between disease severity and growth traits were negative (−0.20 to −0.44) and were also negative between disease severity and survival (−0.28 to −0.51). There were positive genetic correlations between precocity of phase change and growth traits (0.34 to 0.41) and between precocity of phase change and survival (0.29 to 0.37). Our results demonstrate that selection for low disease severity in the first year and for high proportion of adult foliage in the second year are feasible in order to obtain genetic material for sites at risk of infection of both Mycosphaerella leaf disease and Eucalyptus rust.

Mycosphaerella species are well-known causal agents of leaf diseases on many economically and ecologically important plant species. In Uruguay, a relatively large number of Mycosphaerellaceae and Teratosphaeriaceae are found on Eucalyptus, but nothing is known of these fungi on native Myrtaceae. The aim of this study was to identify Mycosphaerellaceae and Teratosphaeriaceae species associated with leaf diseases on native Myrtaceae in Uruguay and to consider whether host jumps by the pathogen from introduced Eucalyptus to native Myrtaceae have occurred. Several native forests throughout the country were surveyed with special attention given to those located close to Eucalyptus plantations. Five species belonging to the Mycosphaerellaceae and Teratosphaeriaceae clades were found on native Myrtaceous trees and three of these had previously been reported on Eucalyptus in Uruguay. Those occurring both on Eucalyptus and native Myrtaceae included Pallidocercospora heimii, Pseudocercospora norchiensis, and Teratosphaeria aurantia. In addition, Mycosphaerella yunnanensis, a species known to occur on Eucalyptus but not previously recorded in Uruguay, was found on leaves of two native Myrtaceous hosts. Because most of these species occur on Eucalyptus in countries other than Uruguay, it appears that they were introduced in this country and have adapted to be able to infect native Myrtaceae. These apparent host jumps have the potential to result in serious disease problems and they should be carefully monitored.

Establishment of alfalfa crops is continuously threatened by seedling diseases caused by soilborne pathogens. The use of plant beneficial bacteria as inoculants is a feasible and environmentally friendly means to control soil pathogens. Identifying effective plant growth-promoting strains to use on local crops under local environmental conditions requires the screening of large collections of native isolates. A collection of 738 rhizospheric fluorescent Pseudomonas isolates was obtained from alfalfa plants from three agroecological regions representative of Uruguayan agricultural systems. The isolates were evaluated for in vitro pathogen inhibition, biosurfactant production, phosphate solubilization and the presence of genes involved in antibiotic synthesis. Isolates with strong in vitro antagonistic activity toward Pythium debaryanum were more abundant in alfalfa plants established in a previously natural ecosystem while biosurfactant producers were less abundant in that location. A subset of isolates was selected for genotypic characterization by rep-PCR using BOX primers. Twenty-four genotypes were defined, sixteen from a single geographical origin and eight composed of isolates from multiple origins. Genotypic profiles correlated well with phenotypic traits. A subset of isolates was assayed to determine their ability to protect alfalfa against P. debaryanum damping-off and to promote plant growth. Five native Pseudomonas isolates showed significant effects on alfalfa by increasing plant biomass and/or protecting from pathogen infection. Plant growth promoting isolates from each location were genotypically similar. Our work contributes to the knowledge of the phenotypic and genotypic diversity of rhizospheric fluorescent pseudomonads of forage legumes and the frequency of plant growth promoting traits associated with this group of bacteria in different agricultural systems.

Streptomyces is a genus of gram-positive bacteria with a mycelial growth habit and the ability to produce spores. Due to their unparalleled ability to produce antibiotics, most of the early research carried out on Streptomyces was antibiotic discovery-driven, with over two thirds of antibiotics used for medical purposes originally isolated from Streptomyces. However, their ubiquity, high capacity of adaptation to different niches and rich secondary metabolite production, make them an invaluable source of solutions in diverse human activities, including medicine, agriculture, industry and toxic waste remotion. In addition to the ability to culture and produce Streptomyces and Streptomyces-derived metabolites, knowledge on how to manipulate natural populations of Streptomyces will likely improve our ability to make environmentally sustainable decisions.

El Eucalyptus globulus es la especie forestal más plantada en el Uruguay, existiendo más de 270.000 hectáreas de plantaciones comerciales. Sin embargo actualmente está siendo sustituido por otras especies, como E. grandis y E. dunnii, debido a los severos daños foliares provocados por Teratosphaeria nubilosa en las plantaciones jóvenes. La continuidad en la plantación de esta especie depende del rápido desarrollo de material genético resistente a la enfermedad. La variación genética de la resistencia del follaje juvenil y la velocidad del cambio de follaje de juvenil a adulto fueron evaluadas en una prueba de progenies de E. globulus naturalmente infectada por T. nubilosa. El material genético incluyó 194 familias de polinización abierta provenientes de dos huertos semilleros pertenecientes al Instituto Nacional de Investigación Agropecuaria (INIA)

Eucalyptus globulus is the most important forest species in Uruguay, with more than 250,000 ha of commercial plantations. Despite its high susceptibility to diseases, production losses caused by foliar diseases have not been properly quantified in this country. This study analyzes the effects of foliar damage on growth and survival using data from a progeny test of E. globulus naturally infected by Teratosphaeria leaf disease and eucalypt rust (Puccinia psidii). The severity of leaf spots and defoliation were quantified 8 months after planting and tree growth and mortality were evaluated 2, 4 and 6 years later. The trial had a high incidence of foliar damage, with a mean leaf spot severity of 28.7% and a mean defoliation of 37%. The greatest impact of foliar damage, both on growth rate and mortality, occurred in the first 2 years after damage was assessed. During this period, leaf spot severity less than 40% and defoliation below 50% did not affect growth, while survival was affected when leaf damage was 70% or greater. By the sixth year both stem growth and survival were affected by severe foliar damage (spotting or defoliation of 80% or more), with a loss of up to 25% in diameter and an accumulated mortality over 70%. It has been established for the first time that under the intensive Uruguayan productive conditions, E. globulus trees can tolerate a relatively high degree of leaf spotting or defoliation but severe foliar damage in the first months can cause considerable production losses, putting at risk the economical viability of this species.

Eucalyptus or guava rust caused by Puccinia psidii is a serious disease of Eucalyptus and other Myrtaceae. In Uruguay, it has been previously found on Eucalyptus globulus and Psidium brasiliensis. Almost nothing is known regarding the occurrence of this pathogen on other Eucalyptus species or native Myrtaceae in that country. In this study, we determined the presence of P. psidii on Eucalyptus species and native Myrtaceae trees in Uruguay and evaluated the pathogenicity of specimens from native myrtaceous hosts on E. globulus and E. grandis. Phylogenetic analyses based on the internal transcribed spacer (ITS) region of the nuclear ribosomal DNA operon were used to confirm pathogen identity. Comparisons of ITS sequences confirmed the identity of P. psidii on Eucalyptus globulus, E. grandis, Myrcianthes pungens, and Myrrhinium atropurpureum var. octandrum. This is the first report of P. psidii on M. atropurpureum var. octandrum. Pathogenicity tests showed that isolates from native Myrtaceae could infect both Eucalyptus species tested, indicating a strong biological relationship between both introduced and native Myrtaceae. This study supplies relevant field data, morphological information, molecular phylogenetic analyses and infection studies that contribute to a better understanding of an important and little studied pathogen.

Symbiotic nitrogen fixation is the main route for sustainable input of nitrogen into ecosystems. Nitrogen fixation in agriculture can be improved by inoculation of legume crops with suitable rhizobia. Knowledge of the biodiversity of rhizobia and of local populations is important for the design of successful inoculation strategies. Soybeans are major nitrogen-fixing crops in many parts of the world. Bradyrhizobial inoculants for soybean are very diverse, yet classification and characterization of strains have long been difficult. Recent genetic characterization methods permit more reliable identification and will improve our knowledge of local populations. Forage legumes form another group of agronomically important legumes. Research and extension policies valorizing rhizobial germplasm diversity and preservation, farmer training for proper inoculant use and legal enforcement of commercial inoculant quality have proved a successful approach to promoting the use of forage legumes while enhancing biological N 2 fixation. It is worth noting that taxonomically important strains may not necessarily be important reference strains for other uses such as legume inoculation and genomics due to specialization of the different fields. This article points out both current knowledge and gaps remaining to be filled for further interaction and improvement of a rhizobial commons.

Species of the Botryosphaeriaceae are important pathogens causing cankers and die-back on many woody plants. In Uruguay, Neofusicoccum eucalyptorum, N. ribis and B. dothidea have previously been associated with stem cankers on plantation-grown Eucalyptus globulus. However, very little is known regarding the occurrence and species diversity of Botryosphaeriaceae in native Myrtaceae forests or what their relationship is to those species infecting Eucalyptus in plantations. The objectives of this study were to identify the Botryosphaeriaceae species present as endophytes or associated with cankers in both introduced and native tree hosts in Uruguay, and to test the pathogenicity of selected isolates obtained from native trees on Eucalyptus. Symptomatic and asymptomatic material was collected countrywide from Eucalyptus plantations and native Myrtaceae trees. Single spore cultures were identified based on conidial morphology and comparisons of DNA sequences of the ITS and EF1-α regions. Six Botryosphaeriaceae species were identified. Botryosphaeria dothidea, N. eucalyptorum and specimens residing in the N. parvum-N. ribis complex were isolated from both introduced Eucalyptus and native Myrtaceae trees, whereas Lasiodiplodia pseudotheobromae was found only on Myrcianthes pungens. Diplodia pseudoseriata sp. nov. and Spencermartinsia uruguayensis sp. nov. are novel species found only on native myrtaceous hosts. Pathogenicity tests showed that isolates obtained from native trees and identified as L. pseudotheobromae, N. eucalyptorum and the N. parvum-N.ribis complex are pathogenic to E. grandis. Interestingly, Lasiodiplodia pseudotheobromae has not previously been found on Eucalyptus in Uruguay and represents a potential threat to this host.

Summary Mycosphaerella leaf diseases represent one of the most important impediments to Eucalyptus plantation forestry. Yet they have been afforded little attention in Uruguay where these trees are an important resource for a growing pulp industry. The objective of this study was to identify species of Mycosphaerellaceae and Teratosphaeriaceae resulting from surveys in all major Eucalyptus growing areas of the country. Species identification was based on morphological characteristics and DNA sequence comparisons for the Internal Transcribed Spacer (ITS) region of the rDNA operon. A total of ten Mycosphaerellaceae and Teratosphaeriaceae were found associated with leaf spots and stem cankers on Eucalyptus. Of these, Mycosphaerella aurantia, M. heimii, M. lateralis, M. scytalidii, Pseudocercospora norchiensis, Teratosphaeria ohnowa and T. pluritubularis are newly recorded in Uruguay. This is also the first report of M. aurantia occurring outside of Australia, and the first record of P. norchiensis and T. pluritubularis in South America. New hosts were identified for Kirramyces gauchensis, M. aurantia, M. marksii, M. lateralis, M. scytalidii, P. norchiensis, T. molleriana, T. ohnowa and T. pluritubularis. Interestingly K. gauchensis, which has been known only as a stem pathogen, was isolated from leaf spots on E. maidenii and E. tereticornis. The large number of Mycosphaerellaceae and Teratosphaeriaceae occurring in Uruguay is disturbing and raises concerns regarding the introduction of new pathogens that could threaten not only Eucalyptus plantations but also native forests.

Neofusicoccum eucalyptorum is a canker-associated fungus apparently highly specialized on Eucalyptus. However, in surveys of the microbial population inhabiting native Myrtaceae in Uruguay, fungal cultures resembling N. eucalyptorum were isolated. The possible occurrence of N. eucalyptorum on hosts other than Eucalyptus prompted further investigation. Several surveys were conducted throughout Uruguay to obtain samples from native forests, focusing primarily on species in the Myrtaceae. Fungal identification was based on morphology and confirmed using comparison sequences for the internal transcribed spacer (ITS) of the rDNA operon. Pathogenicity was evaluated by inoculating plants of a Eucalyptus grandis clone. Morphological and molecular identification confirmed the occurrence of N. eucalyptorum on Blepharocalyx salicifolius, Myrceugenia glaucescens and Myrrhinium atropurpureum var. octandrum. This is the first report of N. eucalyptorum occurring in hosts other than Eucalyptus. Pathogenicity tests confirmed the ability of this species to produce cankers on E. grandis. This study provides information that will assist breeding programmes in attempts to obtain disease-resistant Eucalyptus plantations and it also suggests that a Eucalyptus pathogen could have moved to native trees in Uruguay.

Alfalfa (Medicago sativa) is one of the most important crops used in Uruguay for livestock feeding. Seedling diseases, particularly damping-off, are a critical factor which limits its establishment. Three native Pseudomonas fluorescens strains, UP61.2, UP143.8 and UP148.2, previously isolated from Lotus corniculatus, were evaluated to determine their efficacy as biological control agents for alfalfa seedling diseases in the field. Their compatibility with the alfalfa-Sinorhizobium meliloti symbiosis was also assessed. In growth chamber conditions seed inoculation with Pseudomonas strains did not affect different parameters of alfalfa-rhizobium symbiosis as shown by nodulation rate and shoot dry weight of plants. The presence of the commercial inoculant strains of S. meliloti did not impair colonization by the P. fluorescens and vice versa. In field trials the dynamics of rhizobial rhizospheric populations were not affected by the presence of P. fluorescens. Each P. fluorescens strain successfully colonized alfalfa roots at adequate densities for biocontrol activity. Results showed that P. fluorescens strains provided a 10–13% increase in the number of established plants relative to the control, an intermediate result compared to the fungicide treatment (24%). The alfalfa above-ground biomass was increased by 13% and 15–18% in the presence of the fungicide and P. fluorescens strains, respectively. Therefore, results from this study demonstrated that the three P. fluorescens strains provided effective control against soil-borne pathogens and suggest a potential use in the development of a commercial inoculant to be applied for the control of legume seedling diseases.

Pseudomonas fluorescens strains UP61, UP143 and UP148, isolated from Uruguayan soils, have shown the ability to control soil-borne fungal pathogens that cause damping-off in birdsfoot trefoil. In this communication, we study the effect of these strains on the symbiotic efficiency of rhizobia from commercial inoculants in birdsfoot trefoil, alfalfa and white clover. Shoot dry weights and the rate of nodulation by rhizobia were not modified by the presence of Pseudomonas strains, despite antagonistic activity against rhizobia in vitro. Survival of P. fluorescens UP61 and rhizobia on roots in non-sterile soil were not affected by co-inoculation of the selected forage legumes.

Forage legumes are essential for an efficient animal-based agriculture. This chapter describes microbial-based strategies for improving legume establishment. It focuses on the use of root nodule bacteria for growth promotion and rhizosphere pseudomonads for disease management. The technological system for biological nitrogen fixation deployed in Uruguay is discussed as a model. We analyze the significance of the characterization of rhizobial strains, selection and culture collection, as well as inoculant production and quality control. Knowledge of the biodiversity of introduced and local populations of rhizobia is discussed for the design of successful inoculation strategies. On the other hand, the exploitation of the highly diverse genus Pseudomonas is considered for the biological control of Pythium seedling diseases. By testing large local collections of fluorescent Pseudomonas, strains can be selected with enhanced disease-suppressing and plant growth-promoting abilities to develop bacterial inoculants. The ecology of forage legume microbes is recognized as a key tool for developing sustainable agricultural systems.

The highly diverse genus Pseudomonas contains very effective biocontrol agents that can increase plant growth and improve plant health. Biocontrol characteristics, however, are strain-dependent and cannot be clearly linked to phylogenetic variation. Isolate screening remains essential to find suitable strains, which can be done by testing large local collections for disease suppression and plant-growth promotion exemplified in a case study on forage legumes in Uruguay or by targeted screening for Pseudomonas spp. which produce desirable secondary metabolites, as demonstrated in a case study on cocoyam in Cameroon. In both case studies, access to reference strains from public and private collections was essential for identification, phylogenetic studies and metabolite characterization.

Since the first report of Teratosphaeria nubilosa (Cooke) Crous & U.Braun in Uruguay in 2007, young plantations of Eucalyptus globulus Labill. and E. maidenii F.Muell. have been severely damaged by Mycosphaerella leaf disease. The genetic variation in disease resistance and in the timing of heteroblastic phase change was examined in 194 open-pollinated families of E. globulus and 86 families of E. maidenii growing in a field trial in south-eastern Uruguay, naturally infected by T. nubilosa. Disease severity, precocity of vegetative phase change and tree growth were assessed at 14 months. E. globulus was significantly more susceptible to T. nubilosa than was E. maidenii, presenting higher severity of leaf spots (10.6% and 5.6%, respectively), higher defoliation (31.9% and 22.9%, respectively) and higher crown-damage index (39.1% and 27.4%, respectively). However, the heteroblastic transition began significantly earlier in E. globulus than in E. maidenii, with 34.1% and 2.8% of the trees having some proportion of their crown with adult foliage at 14 months, respectively. Significant individual narrow-sense heritabilities were found in E. globulus for severity of leaf spots (0.40), defoliation (0.24), crown-damage index (0.30) and proportion of adult foliage (0.64). Additive genetic variation in E. maidenii was significant only for defoliation and crown-damage index, with a moderate heritability (0.21 and 0.20, respectively). Although E. maidenii was more resistant to T. nubilosa than was E. globulus, the degree of resistance was not enough to consider this species as an alternative to E. globulus for high-risk disease sites. In addition, the small genetic variability for resistance on the juvenile foliage and the late transition to adult foliage suggested that the chances for early selection in E. maidenii are quite limited. By contrast, the genetic variation in E. globulus clearly indicated that through selection for resistance of the juvenile foliage, and especially by selecting for early phase change, it is possible to obtain genetic stock suitable for sites with high risk of T. nubilosa infection.

Quambalaria eucalypti, a serious pathogen of Eucalyptus, is believed to be native to Australia and introduced into various southern hemisphere countries, including Uruguay. In this study we record the discovery of Q. eucalypti causing leaf lesions on Myrceugenia glaucescens, a tree native to Uruguay. The identity of the pathogen was confirmed using DNA sequence comparisons of the internal transcribed spacer (ITS) region of the rDNA operon, as well as morphological characteristics. This is the first record of the pathogen on a host other than Eucalyptus. It clearly indicates a disturbing example of an alien invasive pathogen having undergone a host-shift from non-native Eucalyptus to a native tree in Uruguay.