Safety assessment of surfactin-producing Bacillus strains and their lipopeptides extracts in vitro and in vivo.

Beneficial Bacillus strains can be administered to livestock as probiotics to improve animal health. Cyclic lipopeptides produced by Bacillus such as surfactins may be responsible for some of the beneficial effects due to their anti-inflammatory and immunomodulatory activity. The aim of the present study was to isolate and evaluate the biocompatibility of native Bacillus spp. strains and their surfactin-like lipopeptides in vitro and in vivo to determine their potential to be used on animals. Biocompatibility of endospore suspensions (108 UFC/mL), and different dilutions (1:10; 1:50; 1:100; 1:500, and 1:1000) of Bacillus lipopeptide extracts containing surfactin was tested on Caco-2 cells by microculture tetrazolium-based colorimetric assay. Genotoxicity was tested on BALB/c mice (n = 6) administered 0.2 mL of endospore suspensions by the bone marrow erythrocyte micronuclei assay. All the isolates tested produced between 26.96 and 239.97 µg mL- 1 of surfactin. The lipopeptide extract (LPE) from isolate MFF1.11 demonstrated significant cytotoxicity in vitro. In contrast, LPE from MFF 2.2; MFF 2.7, TL1.11, TL 2.5, and TC12 had no cytotoxic effect (V% > 70%) on Caco-2 cells, not affecting cell viability signifficantly in most treatments. Similarly, none of the endospore suspensions affected cell viability (V% > 80%). Likewise, endospores did not cause genotoxicity on BALB/c mice. This study was elementary as a first step for a new line of research, since it allowed us to choose the safest isolates to keep working on the search of new potentially probiotic strains destined to production animals to improve their performance and health.

Lactic acid bacteria and Bacillus spp. as fungal biological control agents.

Fungal pathogens are one of the most important agents affecting crop production and food safety, and agrochemical application is one of the main approaches to reduce phytopathogenic fungi contamination in agricultural products. However, excessive and inadequate use can cause environmental damage, human and animal hazard, and increased phytopathogen resistance to fungicides. Biological control using lactic acid bacteria (LAB) and Bacillus spp. is an environmentally friendly strategy for phytopathogenic fungi management. Several molecules produced by these bacteria indeed affect fungal growth and viability in different plant crops. In this article, the activity spectra are reviewed along with the antifungal effect and antifungal compounds produced by LAB (e.g. organic acids, peptides, cyclic dipeptides, fatty acids, and volatile compounds) and Bacillus spp. (e.g. peptides, enzymes, and volatile compounds).

The potential of PGPR and Trichoderma-based bioproducts and resistant cultivars as tools to manage clubroot disease in cruciferous crops.

The objective of this research was to determine the potential use of eco-friendly technologies to reduce the clubroot disease caused by Plasmodiophora brassicae, the main constraint of cruciferous crops worldwide. Two commercial bioproducts were evaluated in susceptible broccoli, one based on the PGPR consortium (Bacillus amyloliquefaciens, Bacillus pumilus, and Agrobacterium radiobacter K84) and the other one based on Trichoderma koningiopsis Th003 (Tricotec® WG). Additionally, the resistant broccoli cv. Monclano® was tested under two concentrations of resting spores (RS) of P. brassicae, 1 × 103 and 1 × 105 RS g-1 of soil. The first phase of evaluations with broccoli was carried out under a greenhouse, while susceptible broccoli, cauliflower, and red cabbage were included in a subsequent field phase. Tebuconazole + Trifloxystrobin mixture and Fluazinam were included as positive controls. The effectiveness of the bioproducts depended on the nature of the biocontrol agent, the concentration of P. brassicae, and the dose of treatment. Tricotec® showed consistent plant growth promotion but no biocontrol effect against clubroot, and the rhizobacteria-based bioproduct significantly reduced the disease in both greenhouse and field experiments. Higher disease severity was observed with the higher dose of Tricotec®. Under field conditions, the rhizobacteria reduced the incidence progress by 26%, 39%, and 57% under high, medium, and low pressure of the pathogen, respectively. However, no reduction of clubroot severity under high pressure of the pathogen was observed. Complete inhibition of club formation in roots was achieved via the fungicide, but a phytotoxic effect was observed under greenhouse conditions. Fungicides reduced the incidence progress of clubroot, but not the severity under high inoculum pressure in the field. The fungicides, the bacterial treatment, and the combination of bioproducts tended to delay the progress of the disease compared with the negative control and Tricotec alone. The resistant broccoli showed a low level of disease under high concentrations of P. brassicae (less than 10% incidence and up to 2% severity). These results suggested the overall potential of commercial tools based on the PGPR consortium and plant resistance to control P. brassicae. The integration of control measures, the role of Trichoderma spp. in P. brassicae-cruciferous pathosystems, and the need to recover highly infested soils will be discussed.

Elicitation of Bacillus cereus-Amazcala (B.c-A) with SiO2 Nanoparticles Improves Its Role as a Plant Growth-Promoting Bacteria (PGPB) in Chili Pepper Plants.

Agriculture needs to decrease the use of agrochemicals due to their high toxicity and adopt new strategies to achieve sustainable food production. Therefore, nanoparticles (NPs) and plant growth-promoting bacteria (PGPB) have been proposed as viable strategies to obtain better crop yields with less environmental impact. Here, we describe the effect of silica nanoparticles (SiO2-NPs) on survival, antioxidant enzymatic activity, phosphate solubilization capacity, and gibberellin production of Bacillus cereus-Amazcala (B.c-A). Moreover, the effect of the co-application of SiO2-NPs and B.c-A on seed germination, physiological characteristics, and antioxidant enzymatic activity of chili pepper plants was investigated under greenhouse conditions. The results indicated that SiO2-NPs at 100 ppm enhanced the role of B.c-A as PGPB by increasing its phosphate solubilization capacity and the production of GA7. Moreover, B.c-A catalase (CAT) and superoxide dismutase (SOD) activities were increased with SiO2-NPs 100 ppm treatment, indicating that SiO2-NPs act as a eustressor, inducing defense-related responses. The co-application of SiO2-NPs 100 ppm and B.c-A improved chili pepper growth. There was an increase in seed germination percentage, plant height, number of leaves, and number and yield of fruits. There was also an increase in CAT and PAL activities in chili pepper plants, indicating that bacteria-NP treatment induces plant immunity.

Ludwig's angina complicated with a massive necrotizing fasciitis caused by an unusual pathogen: Bacillius spp

Ludwig's angina and necrotizing fasciitis are aggressive infectious manifestations that can present in the cervicofacial region. The severity of these entities depends, up to some extent, to the immunological state of the patient. Diabetes mellitus alters the immune system, allowing maxillofacial infections to progress aggressively. Bacillius spp. is a group of typically innocuous bacteria; however, they can unchain relevant infectious pathologies in immunocompromised patients. The purpose of this study is to describe a case of an uncontrolled diabetic patient with a Ludwig's angina which progressed to a massive necrotizing fasciitis caused by bacillius spp. Explaining the severity of this class of infections and the special attention that should be paid to this type of case; Furthermore, when the culture reveals pathogens that are usually not involved in the development of this class of conditions.

In vitro biocontrol of Fusarium oxysporum with antagonistic microorganisms and In vivo effect on Solanum lycopersicum L.

The objective was to evaluate the effects of In vitro biocontrol of Trichoderma spp. and Bacillus spp. against Fusarium oxysporum and its in vivo effect on Solanum lycopersicum plants. The antagonistic effect in vitro of microcapsules of Trichoderma spp. and Bacillus spp. against F. oxysporum, was evaluated by inhibition and antagonism classification, in the in vivo part growth variables in plant and fruit were measured. Trichoderma asperellum showed inhibition of 84.7% and class 1 and Bacillus spp., obtained an inhibition of 83.7% with a positive classification. The canonical correspondence analysis of the fruit, the T1 treatment indicates that the variables that highlight the effect of the treatment are the firmness of the fruit and the Brix. The canonical correspondence analysis of the plant, the height presents a significant difference (P < 0.05), highlighting B. subtilis, Bacillus liquefasciens, T. harzianum and T. asperellum + Bacillus amiloliquefasciens, with 1.05 and 1.1 m compared to the control of 74 cm, also highlighted the number of total fruits in the T. harzianum treatment with an average of 22.6 fruits and root weight in B. subtilis with 37.12 g. Strains of Trichoderma spp. and Bacillus spp. successfully inhibited F. oxysporum also induced growth in the tomato plant.

Comparative Performance of Fungicides and Biocontrol Products in the Management of Fusarium Wilt of Blackberry.

Fusarium wilt of blackberry (FWB) is an emerging disease caused by a Fusarium oxysporum species complex. More than 3,000 ha of blackberry (Rubus spp.) crops have been lost in Mexico since 2011. The objectives of this research were: to evaluate the sensitivity of pathogenic F. oxysporum isolates recovered from symptomatic blackberry plants to fungicides with different modes of action; to assess the potential of these fungicides and plant resistance inducers against FWB in the greenhouse; and to determine the effects of commercial biofungicides and two indigenous strains of Trichoderma spp. on the incidence of FWB. The EC50 values of the fungicides prochloraz, thiabendazole, azoxystrobin, thiophanate-methyl, difenoconazole, triflumizole, and potassium phosphite for six pathogenic F. oxysporum isolates were determined. In a separate experiment, the fungicides acibenzolar-s-methyl (ASM), potassium phosphite, and commercial biofungicides, as well as two soil microbial inoculants and two indigenous Trichoderma strains, were tested for protection against wilt development in blackberry plants in the greenhouse. Prochloraz showed an average sensitivity for EC50 of 0.01 µg ml-1 for the tested F. oxysporum isolates, followed by difenoconazole and thiabendazole. Prochloraz and ASM proved to be the most effective treatments in the greenhouse. In contrast, potassium phosphite was ineffective in both the in vitro and in vivo experiments. The soil bioinoculants MicroSoil, Baktillis, T. koningiopsis, and T. asperellum significantly reduced the incidence of disease in the greenhouse. These results provide evidence for the potential of the various tools as useful components of integrated FWB management in the field.

Dynamics of volatilomes emitted during cross-talking of plant-growth-promoting bacteria and the phytopathogen, Fusarium solani.

The dynamics of volatilomes emitted during the interaction between plant-growth-promoting bacteria (PGPB) and the phytopathogen Fusarium solani were evaluated for 5 days. The first screening was done to evaluate the antagonist activity of volatile compounds emitted by PGPB against F. solani. Volatilomes from 11 PGPB were determined individually and together with F. solani by using solid-phase microextraction coupled to gas-chromatography-mass spectrometry. Isolates of PGPB belonged to the Bacillus genus and inhibited from 18 to 24% the fungal mycelium growth. The isolates also induced morphological alterations of fungal hyphae, like small globular vesicles and the formation of chlamydospores, suggesting a stress mechanism response by the fungus. Volatilome profile showed 49 different compounds that appeared in the bacterial-fungal interaction, such as ketones, sesquiterpenes, monoterpenoids, alkanes, alkenes, carboxylic acids, and fatty acids. Some ketones and alcohols were detected in high abundance only in the interaction PGPB-fungus at 3 and 5 days. Bacillus circulans A19, Bacillus amyloliquefaciens A21, and Bacillus wiedmannii S18 shared a group of emitted alcohols and ketones when they were exposed to F. solani. F. solani produced its own volatilome profile, with the presence of sesquiterpenes, such as α-cubebene and caryophyllene, which increased significantly in co-incubation with the tested bacteria, suggesting chemical communication between them.

Complete Genome Sequence of the Biocontrol Agent Bacillus velezensis UFLA258 and Its Comparison with Related Species: Diversity within the Commons.

In this study, the full genome sequence of Bacillus velezensis strain UFLA258, a biological control agent of plant pathogens was obtained, assembled, and annotated. With a comparative genomics approach, in silico analyses of all complete genomes of B. velezensis and closely related species available in the database were performed. The genome of B. velezensis UFLA258 consisted of a single circular chromosome of 3.95 Mb in length, with a mean GC content of 46.69%. It contained 3,949 genes encoding proteins and 27 RNA genes. Analyses based on Average Nucleotide Identity and Digital DNA-DNA Hybridization and a phylogeny with complete sequences of the rpoB gene confirmed that 19 strains deposited in the database as Bacillus amyloliquefaciens were in fact B. velezensis. In total, 115 genomes were analyzed and taxonomically classified as follows: 105 were B. velezensis, 9 were B. amyloliquefaciens, and 1 was Bacillus siamensis. Although these species are phylogenetically close, the combined analyses of several genomic characteristics, such as the presence of biosynthetic genes encoding secondary metabolites, CRISPr/Cas arrays, Average Nucleotide Identity and Digital DNA-DNA Hybridization, and other information on the strains, including isolation source, allowed their unequivocal classification. This genomic analysis expands our knowledge about the closely related species, B. velezensis, B. amyloliquefaciens, and B. siamensis, with emphasis on their taxonomical status.

Screening of bacterial extracellular xylanase producers with potential for cellulose pulp biobleaching

In this study, two hundred fifty-seven bacterial isolates from a suppressive soil library were screened to study their secretion of alkali-thermostable xylanases for potential use in cellulose pulp biobleaching. Xylanase activity was evaluated in solid and liquid media using xylan as the carbon source. Isolates were initially evaluated for the degradation of xylan in solid media by the congo red test. Selected strains were evaluated in liquid media for enzymatic activity and determination of total protein concentration using a crude protein extract (CPE). An isolate identified as Bacillus species TC-DT13 produced the highest amount of xylanase (1808 U mL-1). The isolate was active and stable at 70°C and pH 9.0, conditions which are necessary for the paper industry. This isolate can grow and produce xylanase in medium containing wheat fiber as a substrate. The CPE of this isolate was used in preliminary testing on cellulose pulp bleaching; enzyme treatment of the pulp resulted in a 5% increase of whiteness.

Screening of bacterial extracellular xylanase producers with potential for cellulose pulp biobleaching

In this study, two hundred fifty-seven bacterial isolates from a suppressive soil library were screened to study their secretion of alkali-thermostable xylanases for potential use in cellulose pulp biobleaching. Xylanase activity was evaluated in solid and liquid media using xylan as the carbon source. Isolates were initially evaluated for the degradation of xylan in solid media by the congo red test. Selected strains were evaluated in liquid media for enzymatic activity and determination of total protein concentration using a crude protein extract (CPE). An isolate identified as Bacillus species TC-DT13 produced the highest amount of xylanase (1808 U mL-1). The isolate was active and stable at 70°C and pH 9.0, conditions which are necessary for the paper industry. This isolate can grow and produce xylanase in medium containing wheat fiber as a substrate. The CPE of this isolate was used in preliminary testing on cellulose pulp bleaching; enzyme treatment of the pulp resulted in a 5% increase of whiteness.(AU)

Prevalence and etiology of mastitis in dairy cattle in El Oro Province, Ecuador.

This study described the occurrence of clinical and subclinical forms of mastitis in 250 cattle from 5 dairy farms around the cities of Santa Rosa and Machala, El Oro Province, Ecuador. Clinical mastitis (CM) was determined based on obvious changes in milk (mild), signs of inflammation in the udder (moderate), and/or generalized clinical symptoms (severe). Subclinical mastitis (SCM) was assessed using the California mastitis test. CM and SCM were detected in 30 (12.0%) and 150 (60%) of the 250 tested cattle, respectively. Prevalence at the udder quarter level was 57.7% (577/1,000), which was higher among forequarters (369/577; 63.9%) than hindquarters. Of the 577 mastitic milk samples subjected to microbiological analysis, 35 were excluded due to contamination and 20 tested negative. Identification of bacterial isolates revealed that 33.3% of the 93 CM samples contained coliforms, 25.8% coagulase-positive staphylococci, 20.4% coagulase-negative staphylococci (CNS), 9.7% streptococci, 7.5% Bacillus spp., and 3.2% Klebsiella spp. Bacterial profiling of the 429 SCM milk samples showed that 55.4% contained CNS, 22.1% Bacillus spp., 9.3% streptococci, and 6.1% coagulase-positive staphylococci. In vitro antibiotic susceptibility testing of the obtained isolates indicated that all were susceptible to amoxicillin, ampicillin, cefotaxime, enrofloxacin, sulfamethoxazole-trimethoprim, gentamicin, and neomycin. No multidrug-resistant strains were observed.

Diversity and enzymatic potentialities of Bacillus sp. strains isolated from a polluted freshwater ecosystem in Cuba.

Genotypic and phenotypic characterization of Bacillus spp. from polluted freshwater has been poorly addressed. The objective of this research was to determine the diversity and enzymatic potentialities of Bacillus spp. strains isolated from the Almendares River. Bacilli strains from a polluted river were characterized by considering the production of extracellular enzymes using API ZYM. 14 strains were selected and identified using 16S rRNA, gyrB and aroE genes. Genotypic diversity of the Bacillus spp. strains was evaluated using pulsed field gel electrophoresis. Furthermore, the presence of genetic determinants of potential virulence toxins of the Bacillus cereus group and proteinaceous crystal inclusions of Bacillus thuringiensis was determined. 10 strains were identified as B. thuringiensis, two as Bacillus megaterium, one as Bacillus pumilus and one as Bacillus subtilis. Most strains produced proteases, amylases, phosphatases, esterases, aminopeptidases and glucanases, which reflect the abundance of biopolymeric matter in Almendares River. Comparison of the typing results revealed a spatio-temporal distribution among B. thuringiensis strains along the river. The results of the present study highlight the genotypic and phenotypic diversity of Bacillus spp. strains from a polluted river, which contributes to the knowledge of genetic diversity of Bacilli from tropical polluted freshwater ecosystems.

Bacillus spp: una alternativa para la promoción vegetal por dos caminos enzimáticos / Bacillus spp: An alternative for plant promotion by two enzymatic pathways

Resumen Objetivo. Se realizó una revisión sobre las características de las fitasas y nitrogenasas de Bacillus spp. y sus opciones de uso como alternativa biofertilizante. El género Bacillus es secretor de proteínas y metabolitos eficientes para el control de plagas y enfermedades, promueve el crecimiento vegetal a través de la solubilización de fósforo y la producción de reguladores de crecimiento como el ácido indol acético; así mismo participa en la fijación de nitrógeno cuando hace parte de consorcios microbianos. Como biofertilizante es una opción amigable para el suelo y el ambiente que da respuesta a la necesidad de implementar la agricultura sostenible.

Abstract Objective. We conducted a review of the characteristics of the phytase and nitrogenase of Bacillus sp. and their potential use as an alternative biological fertilizer. The genus Bacillus is an efficient secretor of proteins and metabolites; to control pests and diseases, promote plant growth through the solubilization of phosphorus and production of growth regulators as acetic indole. Likewise it is involved in nitrogen fixation when it is part of microbial consortia. As biological fertilizer is a friendly option for the soil and the environment that responds to the need to implement sustainable agriculture.

Biodegradation of Diesel, Crude Oil and Spent Lubricating Oil by Soil Isolates of Bacillus spp.

Two species of Bacillus, B. thuringiensis B3 and B. cereus B6, isolated from crude oil-contaminated sites in Ecuador, were tested for their capability in degrading polycyclic aromatic hydrocarbons (PAHs) in diesel (shake-flask), and to remove total petroleum hydrocarbons (TPHs) from crude oil- or spent lubricating oil-polluted soils (plot-scale). TPHs and PAHs were analyzed by Gas chromatography-Flame ionization detector (GC-FID) and High performance liquid chromatography (HPLC), respectively. Degradation percentages of PAHs by strain B6 were in the range of 11-83 after 30 days. A mixed culture of both the strains removed 84% and 28% of TPHs from crude oil- and spent lubricating oil-polluted soils, respectively. Reduction in the abundance of total n-alkane fractions (C8-C40) of spent lubricating oil was 94%, which was 18% higher than the control. Our results clearly indicate that the selected strains have great potential in degrading petroleum hydrocarbons at both laboratory- and field-scales.

Microbial degradation of total petroleum hydrocarbons in crude oil: a field-scale study at the low-land rainforest of Ecuador.

A field-level feasibility study was conducted to determine total petroleum hydrocarbon (TPH)-degrading potential of two bacterial strains, Bacillus thuringiensis B3 and B. cereus B6, and two fungi, Geomyces pannorum HR and Geomyces sp. strain HV, all soil isolates obtained from an oil field located in north-east region of Ecuador. Crude oil-treated soil samples contained in wooden boxes received a mixture of all the four microorganisms and were incubated for 90 days in an open low-land area of Amazon rainforest. The percent removal of TPHs in soil samples that received the mixed microbial inoculum was 87.45, indicating the great potential of the soil isolates in field-scale removal of crude oil. The TPHs-degrading efficiency was verified by determining the toxicity of residues, remained in soil after biodegradation, toward viability of Artemia salina or seed germination and plant growth of cowpea. Our results clearly suggest that the selected soil isolates of bacteria and fungi could be effectively used for large-scale bioremediation of sites contaminated with crude oil.

Gram-Positive Bacteria with Probiotic Potential for the Apis mellifera L. Honey Bee: The Experience in the Northwest of Argentina.

Apis mellifera L. is one of the most important natural pollinators of significant crops and flowers around the world. It can be affected by different types of illnesses: american foulbrood, nosemosis, varroasis, viruses, among others. Such infections mainly cause a reduction in honey production and in extreme situations, the death of the colony. Argentina is the world's second largest honey exporter and the third largest honey producer, after China and Turkey. Given both the prominence of the honey bee in nature and the economic importance of apiculture in Argentina and the world, it is crucial to develop efficient and sustainable strategies to control honey bee diseases and to improve bee colony health. Gram-positive bacteria, such as lactic acid bacteria, mainly Lactobacillus, and Bacillus spp. are promising options. In the Northwest of Argentina, several Lactobacillus and Bacillus strains from the honey bee gut and honey were isolated by our research group and characterized by using in vitro tests. Two strains were selected because of their potential probiotic properties: Lactobacillus johnsonii CRL1647 and Bacillus subtilis subsp. subtilis Mori2. Under independent trials with both experimental and commercial hives, it was determined that each strain was able to elicit probiotic effects on bee colonies reared in the northwestern region of Argentina. One result was the increase in egg-laying by the queen which therefore produced an increase in bee number and, consequently, a higher honey yield. Moreover, the beneficial bacteria reduced the incidence of two important bee diseases: nosemosis and varroosis. These results are promising and extend the horizon of probiotic bacteria to the insect world, serving beekeepers worldwide as a natural tool that they can administer as is, or combine with other disease-controlling methods.

Effect of mixed-Bacillus spp isolated from pustulose ark Anadara tuberculosa on growth, survival, viral prevalence and immune-related gene expression in shrimp Litopenaeus vannamei.

The widespread overuse of antibiotics in aquaculture has led to the emergence of antibiotic-resistance shrimp pathogens, the negative impact on shrimp gut microbiota, and the presence of antimicrobial residues in aquaculture products, with negative consequences on human health. Alternatively, probiotics have positive effects on immunological responses and productive performance of aquatic animals. In this study, three probiotic bacteria, (Bacillus licheniformis MAt32, B. subtilis MAt43 and B. subtilis subsp. subtilis GAtB1), isolated from the Anadara tuberculosa were included in diets for juvenile shrimp, Litopenaeus vannamei, to evaluate their effects on growth, survival, disease prevalence, and immune-related gene expression. Shrimp naturally infected with WSSV and IHHNV were fed with the basal diet (control, T1) and diets supplemented with four levels of bacilli probiotic mix (1:1:1) at final concentration of (T2) 1 × 106, (T3) 2 × 106, (T4) 4 × 106, and (T5) 6 × 106 CFU g-1 of feed. The specific growth rate of shrimp was significantly higher in T2 than in T1 (control) treatment, and the final growth as well as the survival were similar among treated groups. The prevalence of WSSV and IHHNV infected shrimp was reduced in T2 and T4 treatments, respectively, compared with control. The mRNA expression of proPO gene was higher in treatment T4 than control. The LvToll1 gene was significantly up-regulated in treatments T4 and T5 compared to control. The SOD gene was up-regulated in treatment T5 compared to control. In contrast, the mRNA expression of the Hsp70 gene was down-regulated in treatments T4 and T5 respect to control, and the TGase gene remained unaffected by the level of bacillus probiotic mix. As conclusion, the bacilli probiotic mix (Bacillus spp.) enhanced immune-related gene expression in WSSV and IHHNV naturally infected shrimp. This is the first report of probiotic potential of bacteria isolated from A. tuberculosa on the immune response and viral prevalence in shrimp Litopenaeus vannamei.

Bacteria as growth-promoting agents for citrus rootstocks.

The microbial community plays an essential role in maintaining the ecological balance of soils. Interactions between microorganisms and plants have a major influence on the nutrition and health of the latter, and growth-promoting rhizobacteria can be used to improve plant development through a wide range of mechanisms. Therefore, the objective of the present study was to evaluate bacteria as growth-promoting agents for citrus rootstocks. A total of 30 bacterial isolates (11 of Bacillus spp., 11 actinobacteria, and 8 lactic acid bacteria) were evaluated in vitro for indoleacetic acid production, phosphate solubilization, and nitrogen (N) fixation. In vivo testing consisted of growth promotion trials of the bacterial isolates that yielded the best results on in vitro tests with three rootstocks: Swingle citrumelo [Citrus×paradisi Macfad cv. Duncan×Poncirus trifoliata (L.) Raf.], Sunki mandarin (Citrus sunki Hort. ex Tan), and rangpur (Citrus×limonia Osbeck). The parameters of interest were height, number of leaves, stem diameter, shoot and root dry mass, and total dry mass at 150days after germination. The results showed that most bacterial isolates were capable of IAA production. Only one lactic acid bacterium isolate (BL06) solubilized phosphate, with a high solubilization index (PSI>3). In the actinobacteria group, isolates ACT01 (PSI=2.09) and ACT07 (PSI=2.01) exhibited moderate phosphate-solubilizing properties. Of the Bacillus spp. isolates, only CPMO6 and BM17 solubilized phosphate. The bacterial isolates that most fixated nitrogen were BM17, ACT11, and BL24. In the present study, some bacteria were able to promote growth of citrus rootstocks; however, this response was dependent on plant genotype and isolate. Bacillus spp. BM16 and CPMO4 were able to promote growth of Swingle citrumelo. In Sunki mandarin plants, the best treatment results were obtained with BM17 (Bacillus sp.) and ACT11 (actinobacteria). For Rangpur lime rootstock, only BM05 (Bacillus sp.) was able to promote increase in two parameters assessed, height and number of leaves. When the bacterial isolates were used in mixture there was not promoted growth of plants on rootstocks. This fact may be associated with the different mechanisms of action of each bacteria involved or with the presence of competition among the microorganisms of the mixture.

Isolation of Mexican Bacillus Species and Their Effects in Promoting Growth of Chili Pepper (Capsicum annuum L. cv Jalapeño).

The purpose of this work was to isolate and identify native bacteria from plants collected in the State of Yucatán, México with the ability to promote growth of chili pepper (Capsicum annuum L. cv Jalapeño). We identified nine bacterial isolates that belong to five species of Bacillus (i.e. Bacillus subtilis, B. flexus, B. cereus, B. megaterium and B. endophyticus) that produced indoleacetic acid (4.0-24.3 µg/mL) with solubilization index of 1.3-1.6. All the bacterial isolates were evaluated based on their ability to promote growth of chili pepper. Plants inoculated with B. subtilis ITC-N67 showed an increase in stem diameter and root volume, whereas inoculation with B. cereus ITC-BL18 increased the number of flower buds, fresh biomass of roots and total fresh biomass. Conversely, B. flexus ITC-P4 and B. flexus ITC-P22 showed deleterious effect on root volume and total biomass. In summary, our data showed that native B. cereus TC-BL18 and B. subtilis ITC-N67 have potential to be used as growth promoting microorganism for chili pepper, particularly in the state of Yucatán, México.