Characterization of methyl parathion degradation by a Burkholderia zhejiangensis strain, CEIB S4-3, isolated from agricultural soils.

Through the use of an enrichment technique, we isolated from the agricultural soils of Morelos in central México a strain of Burkholderia zhejiangensis identified as CEIB S4-3, it's could use the pesticide methyl parathion (MP) as the only source of carbon and degrade completely p-nitrophenol (PNP). For more efficient MP and PNP degradation by the CEIB S4-3 strain, the absence of an extra carbon source, a large inoculum and an MP concentration up to 50 mg/l are required. Sequence and annotation analysis of the draft genome, showed presence of mpd functional gene, which was expressed and its activity on the MP was confirmed. Additionally, the genes coding for enzymes in the benzoquinone pathway (conducted by Gram-negative bacteria) and the benzenotriol pathway (conducted by Gram-positive bacteria) were found, which was corroborated by identification of intermediary metabolites by HPLC. Thus, we propose that B. zhejiangensis CEIB S4-3 uses both degradation pathways.

Impact of Meyerozyma guilliermondii isolated from chickens against Eimeria sp. protozoan, an in vitro analysis.

BACKGROUND: Avian coccidiosis is a disease caused worldwide by several species of parasite Eimeria that causes significant economic losses. This disease affects chickens development and production, that most of times is controlled with anticoccidial drugs. Although efforts have been made to address this disease, they have been made to control Eimeria sporozoites, although enteric stages are often vulnerable, however; the parasite oocyst remains a problem that must be controlled, as it has a resistant structure that facilitates dispersion. Despite some commercial products based on chemical compounds have been developed as disinfectants that destroy oocysts, the solution of the problem remains to be solved. RESULTS: In this work, we assessed in vitro anticoccidial activity of a compound(s) secreted by yeast isolated in oocysts suspension from infected chickens. The yeast was molecularly identified as Meyerozyma guilliermondii, and its anticoccidial activity against Eimeria tenella oocysts was assessed. Here, we report the damage to oocysts walls caused by M. guilliermondii culture, supernatant, supernatant extract and intracellular proteins. In all cases, a significant decreased of oocysts was observed. CONCLUSIONS: The yeast Meyerozyma guilliermondii secretes a compound with anticoccidial activity and also has a compound of protein nature that damages the resistant structure of oocyst, showing the potential of this yeast and its products as a feasible method of coccidiosis control.

Crude Extracts of Pleurotus Spp. and the Presence of Their Proteins in the Ovicidal and Larvicidal Activity of Haemonchus contortus.

Mushrooms of the genus Pleurotus have shown nematophagous activity as it produces many chemical compounds and enzymes affecting parasitic nematodes. This study aimed to extract the inhibitory activity of the five strains of the fungus Pleurotus spp. It was evaluated against eggs and larvae of Haemonchus contortus. The extract of P. ostreatus obtained the highest level of inhibition of eggs at 97.6% (1341 µg/mL) followed by P. pulmonarius (EPP) at 81.2% (774 µg/mL). The extract selected for evaluation against larvae was P. pulmonarius, showing no effect for L3 larvae, but for L4 larvae an immobility effect of 56.93% was observed at 900 µg/mL. The protein profile showed the presence of 23 protein bands in the extract. The crude extract of P. pulmonarius showed degradation of tissues both inside the eggs and larvae L1. Metabolites produced by Pleurotus mushrooms can consider using in agriculture sustainable by utilizing in producing of ovicidal and larvicidal against H. contortus instead of chemical compounds.

Evaluation and Characterization of the Insecticidal Activity and Synergistic Effects of Different GroEL Proteins from Bacteria Associated with Entomopathogenic Nematodes on Galleria mellonella.

GroEL is a chaperonin that helps other proteins fold correctly. However, alternative activities, such as acting as an insect toxin, have also been discovered. This work evaluates the chaperonin and insecticidal activity of different GroEL proteins from entomopathogenic nematodes on G. mellonella. The ability to synergize with the ExoA toxin of Pseudomonas aeruginosa was also investigated. The GroELXn protein showed the highest insecticidal activity among the different GroELs. In addition, it was able to significantly activate the phenoloxidase system of the target insects. This could tell us about the mechanism by which it exerts its toxicity on insects. GroEL proteins can enhance the toxic activity of the ExoA toxin, which could be related to its chaperonin activity. However, there is a significant difference in the synergistic effect that is more related to its alternative activity as an insecticidal toxin.

Isolation of the opdE gene that encodes for a new hydrolase of Enterobacter sp. capable of degrading organophosphorus pesticides.

Microbial enzymes that can hydrolyze organophosphorus compounds have been isolated, identified and characterized from different microbial species in order to use them in biodegradation of organophosphorus compounds. We isolated a bacterial strain Cons002 from an agricultural soil bacterial consortium, which can hydrolyze methyl-parathion (MP) and other organophosphate pesticides. HPLC analysis showed that strain Cons002 is capable of degrading pesticides MP, parathion and phorate. Pulsed-field gel electrophoresis and 16S rRNA amplification were performed for strain characterization and identification, respectively, showing that the strain Cons002 is related to the genus Enterobacter sp. which has a single chromosome of 4.6 Mb and has no plasmids. Genomic library was constructed from DNA of Enterobacter sp. Cons002. A gene called opdE (Organophosphate Degradation from Enterobacter) consists of 753 bp and encodes a protein of 25 kDa, which was isolated using activity methods. This gene opdE had no similarity to any genes reported to degrade organophosphates. When kanamycin-resistance cassette was placed in the gene opdE, hydrolase activity was suppressed and Enterobacter sp. Cons002 had no growth with MP as a nutrients source.

Mexican Strains of Anaplasma marginale: A First Comparative Genomics and Phylogeographic Analysis.

The One Health approach looks after animal welfare and demands constant monitoring of the strains that circulate globally to prevent outbreaks. Anaplasma marginale is the etiologic agent of bovine anaplasmosis and is endemic worldwide. This study aimed to analyze, for the first time, the genetic diversity of seven Mexican strains of A. marginale and their relationship with other strains reported. The main features of A. marginale were obtained by characterizing all 24 genomes reported so far. Genetic diversity and phylogeography were analyzed by characterizing the msp1a gene and 5'-UTR microsatellite sequences and constructing a phylogenetic tree with 540 concatenated genes of the core genome. The Mexican strains show 15 different repeat sequences in six MSP1a structures and have phylogeographic relationships with strains from North America, South America, and Asia, which confirms they are highly variable. Based on our results, we encourage the performance of genome sequencing of A. marginale strains to obtain a high assembly level of molecular markers and the performance of extensive phylogeographic analysis. Undoubtedly, genomic surveillance helps build a picture of how a pathogen changes and evolves in geographical regions. However, we cannot discard the study of relationships pathogens establish with ticks and how they have co-evolved to establish themselves as a successful transmission system.

Molecular Characterization of pBOq-IncQ and pBOq-95LK Plasmids of Escherichia coli BOq 01, a New Isolated Strain from Poultry Farming, Involved in Antibiotic Resistance.

The increase in antimicrobial resistance has raised questions about how to use these drugs safely, especially in veterinary medicine, animal nutrition, and agriculture. Escherichia coli is an important human and animal pathogen that frequently contains plasmids carrying antibiotic resistance genes. Extra chromosomal elements are required for various functions or conditions in microorganisms. Several phage-like plasmids have been identified, which are important in antibiotic resistance. In this work, the molecular characterization of the pBOq-IncQ (4.5 kb) and pBOq-95LK (95 kb) plasmids found in the E. coli strain BOq 01, a multidrug resistant bacteria isolated from a poultry farm, are considered. Plasmid pBOq-IncQ belongs to the incQ incompatibility plasmid family and is involved in sulfonamide resistance. Plasmid pBOq-95LK is a lytic phage-like plasmid that is involved in the lysis of the E. coli BOq 01 strain and carries a bleomycin resistance gene and a strain cured of this plasmid shows bleomycin sensitivity. Induction of the lytic cycle indicates that this phage-like plasmid is an active phage. This type of plasmid has been reported to acquire genes such as mcr-1, which codes for colistin resistance and bacterial persistence and is a significant public health threat. A genome comparison, a pangenomic and phylogenomic analysis with other phage-like plasmids reported in the literature were performed to understand better the evolution of this kind of plasmid in bacteria and its potential importance in antibiotic resistance.

Comparative Genomics and Pathogenicity Analysis of Two Bacterial Symbionts of Entomopathogenic Nematodes: The Role of the GroEL Protein in Virulence.

Bacteria of the genera Xenorhabdus and Photorhabdus are symbionts of entomopathogenic nematodes. Despite their close phylogenetic relationship, they show differences in their pathogenicity and virulence mechanisms in target insects. These differences were explored by the analysis of the pangenome, as it provides a framework for characterizing and defining the gene repertoire. We performed the first pangenome analysis of 91 strains of Xenorhabdus and Photorhabdus; the analysis showed that the Photorhabdus genus has a higher number of genes associated with pathogenicity. However, biological tests showed that whole cells of X. nematophila SC 0516 were more virulent than those of P. luminescens HIM3 when both were injected into G. mellonella larvae. In addition, we cloned and expressed the GroEL proteins of both bacteria, as this protein has been previously indicated to show insecticidal activity in the genus Xenorhabdus. Among these proteins, Cpn60-Xn was found to be the most toxic at all concentrations tested, with an LC50 value of 102.34 ng/larva. Sequence analysis suggested that the Cpn60-Xn toxin was homologous to Cpn60-Pl; however, Cpn60-Xn contained thirty-five differentially substituted amino acid residues that could be responsible for its insecticidal activity.

High-throughput transcriptome sequencing and comparative analysis of Escherichia coli and Schizosaccharomyces pombe in respiratory and fermentative growth.

In spite of increased complexity in eukaryotes compared to prokaryotes, several basic metabolic and regulatory processes are conserved. Here we explored analogies in the eubacteria Escherichia coli and the unicellular fission yeast Schizosaccharomyces pombe transcriptomes under two carbon sources: 2% glucose; or a mix of 2% glycerol and 0.2% sodium acetate using the same growth media and growth phase. Overall, twelve RNA-seq libraries were constructed. A total of 593 and 860 genes were detected as differentially expressed for E. coli and S. pombe, respectively, with a log2 of the Fold Change ≥ 1 and False Discovery Rate ≤ 0.05. In aerobic glycolysis, most of the expressed genes were associated with cell proliferation in both organisms, including amino acid metabolism and glycolysis. In contrast in glycerol/acetate condition, genes related to flagellar assembly and membrane proteins were differentially expressed such as the general transcription factors fliA, flhD, flhC, and flagellum assembly genes were detected in E. coli, whereas in S. pombe genes for hexose transporters, integral membrane proteins, galactose metabolism, and ncRNAs related to cellular stress were overexpressed. In general, our study shows that a conserved "foraging behavior" response is observed in these eukaryotic and eubacterial organisms in gluconeogenic carbon sources.

Ornithine Lipids in Burkholderia spp. Pathogenicity.

The genus Burkholderia sensu lato is composed of a diverse and metabolically versatile group of bacterial species. One characteristic thought to be unique for the genus Burkholderia is the presence of two forms each (with and without 2-hydroxylation) of the membrane lipids phosphatidylethanolamine (PE) and ornithine lipids (OLs). Here, we show that only Burkholderia sensu stricto strains constitutively form OLs, whereas all other analyzed strains belonging to the Burkholderia sensu lato group constitutively form the two forms of PE, but no OLs. We selected two model bacteria to study the function of OL in Burkholderia sensu lato: (1) Burkholderia cenocepacia wild-type which constitutively forms OLs and its mutant deficient in the formation of OLs and (2) Robbsia andropogonis (formerly Burkholderia andropogonis) which does not form OL constitutively, and a derived strain constitutively forming OLs. Both were characterized under free-living conditions and during pathogenic interactions with their respective hosts. The absence of OLs in B. cenocepacia slightly affected bacterial growth under specific abiotic stress conditions such as high temperature and low pH. B. cenocepacia lacking OLs caused lower mortality in Galleria mellonella larvae while R. andropogonis constitutively forming OLs triggers an increased formation of reactive oxygen species immediately after infection of maize leaves, suggesting that OLs can have an important role during the activation of the innate immune response of eukaryotes.

Whole-Genome Sequencing of Mexican Strains of Anaplasma marginale: An Approach to the Causal Agent of Bovine Anaplasmosis.

Anaplasma marginale is the main etiologic agent of bovine anaplasmosis, and it is extensively distributed worldwide. We have previously reported the first genome sequence of a Mexican strain of A. marginale (Mex-01-001-01). In this work, we report the genomic analysis of one strain from Hidalgo (MEX-14-010-01), one from Morelos (MEX-17-017-01), and two strains from Veracruz (MEX-30-184-02 and MEX-30-193-01). We found that the genome average size is 1.16-1.17 Mbp with a GC content close to 49.80%. The genomic comparison reveals that most of the A. marginale genomes are highly conserved and the phylogeny showed that Mexican strains cluster with Brazilian strains. The genomic information contained in the four draft genomes of A. marginale from Mexico will contribute to understanding the molecular landscape of this pathogen.

The Polycyclic Aromatic Hydrocarbon (PAH) degradation activities and genome analysis of a novel strain Stenotrophomonas sp. Pemsol isolated from Mexico.

BACKGROUND: Stenotrophomonas are ubiquitous gram-negative bacteria, which can survive in a wide range of environments. They can use many substances for their growth and are known to be intrinsically resistant to many antimicrobial agents. They have been tested for biotechnological applications, bioremediation, and production of antimicrobial agents. METHOD: Stenotrophomonas sp. Pemsol was isolated from a crude oil contaminated soil. The capability of this isolate to tolerate and degrade polycyclic aromatic hydrocarbons (PAH) such as anthraquinone, biphenyl, naphthalene, phenanthrene, phenanthridine, and xylene was evaluated in Bushnell Hass medium containing PAHs as the sole carbon sources. The metabolites formed after 30-day degradation of naphthalene by Pemsol were analyzed using Fourier Transform Infra-red Spectroscopic (FTIR), Ultra-Performance Liquid Chromatography-Mass Spectrometry (UPLC-MS) and Gas Chromatography-Mass Spectrometry (GC-MS). The genome of Pemsol was also sequenced and analyzed. RESULTS: Anthraquinone, biphenyl, naphthalene, phenanthrene, and phenanthridine except xylene can be used as sole carbon sources for Pemsol's growth in Bushnell Hass medium. The degradation of naphthalene at a concentration of 1 mg/mL within 30 days was tested. A newly formed catechol peak and the disappearance of naphthalene peak detected on the UPLC-MS, and GC-MS analyses spectra respectively confirmed the complete degradation of naphthalene. Pemsol does not produce biosurfactant and neither bio-emulsify PAHs. The whole genome was sequenced and assembled into one scaffold with a length of 4,373,402 bp. A total of 145 genes involved in the degradation of PAHs were found in its genome, some of which are Pemsol-specific as compared with other 11 Stenotrophomonas genomes. Most specific genes are located on the genomic islands. Stenotrophomonas sp. Pemsol's possession of few genes that are associated with bio-emulsification gives the genetic basis for its inability to bio-emulsify PAH. A possible degradation pathway for naphthalene in Pemsol was proposed following the analysis of Pemsol's genome. ANI and GGDH analysis indicated that Pemsol is likely a new species of Stenotrophomonas. It is the first report on a complete genome sequence analysis of a PAH-degrading Stenotrophomonas. Stenotrophomonas sp. Pemsol possesses features that make it a good bacterium for genetic engineering and will be an excellent tool for the remediation of crude oil or PAH-contaminated soil.

Assessing the Pathogenicity of Two Bacteria Isolated from the Entomopathogenic Nematode Heterorhabditis indica against Galleria mellonella and Some Pest Insects.

The entomopathogenic nematodes Heterorhabditis are parasites of insects and are associated with mutualist symbiosis enterobacteria of the genus Photorhabdus; these bacteria are lethal to their host insects. Heterorhabditis indica MOR03 was isolated from sugarcane soil in Morelos state, Mexico. The molecular identification of the nematode was confirmed using sequences of the ITS1-5.8S-ITS2 region and the D2/D3 expansion segment of the 28S rRNA gene. In addition, two bacteria HIM3 and NA04 strains were isolated from the entomopathogenic nematode. The genomes of both bacteria were sequenced and assembled de novo. Phylogenetic analysis was confirmed by concatenated gene sequence datasets as Photorhabdus luminescens HIM3 (16S rRNA, 23S rRNA, dnaN, gyrA, and gyrB genes) and Pseudomonas aeruginosa NA04 (16S rRNA, 23S rRNA and gyrB genes). H. indica MOR03 infects Galleria mellonella, Tenebrio molitor, Heliothis subflexa, and Diatraea magnifactella larvae with LC50 values of 1.4, 23.5, 13.7, and 21.7 IJs/cm², respectively, at 48 h. These bacteria are pathogenic to various insects and have high injectable insecticide activity at 24 h.

Draft Genome Sequences of Anaplasma marginale Strains MEX-15-099-01 and MEX-31-096-01, Two Mexican Isolates with Different Degrees of Virulence.

Anaplasma marginale is an intraerythrocytic bacterium that causes bovine anaplasmosis and is endemic in Mexico. In this work, we report two draft genome sequences of Mexican isolates from different geographical regions and with different degrees of virulence.

Insect Hsp90 Chaperone Assists Bacillus thuringiensis Cry Toxicity by Enhancing Protoxin Binding to the Receptor and by Protecting Protoxin from Gut Protease Degradation.

Bacillus thuringiensis Cry proteins are pore-forming insecticidal toxins with specificity against different crop pests and insect vectors of human diseases. Previous work suggested that the insect host Hsp90 chaperone could be involved in Cry toxin action. Here, we show that the interaction of Cry toxins with insect Hsp90 constitutes a positive loop to enhance the performance of these toxins. Plutella xylostella Hsp90 (PxHsp90) greatly enhanced Cry1Ab or Cry1Ac toxicity when fed together to P. xylostella larvae and also in the less susceptible Spodoptera frugiperda larvae. PxHsp90 bound Cry1Ab and Cry1Ac protoxins in an ATP- and chaperone activity-dependent interaction. The chaperone Hsp90 participates in the correct folding of proteins and may suppress mutations of some client proteins, and we show here that PxHsp90 recovered the toxicity of the Cry1AbG439D protoxin affected in receptor binding, in contrast to the Cry1AbR99E or Cry1AbE129K mutant, affected in oligomerization or membrane insertion, respectively, which showed a slight toxicity improvement. Specifically, PxHsp90 enhanced the binding of Cry1AbG439D protoxin to the cadherin receptor. Furthermore, PxHsp90 protected Cry1A protoxins from degradation by insect midgut proteases. Our data show that PxHsp90 assists Cry1A proteins by enhancing their binding to the receptor and by protecting Cry protoxin from gut protease degradation. Finally, we show that the insect cochaperone protein PxHsp70 also increases the toxicity of Cry1Ac in P. xylostella larvae, in contrast to a bacterial GroEL chaperone, which had a marginal effect, indicating that the use of insect chaperones along with Cry toxins could have important biotechnological applications for the improvement of Cry insecticidal activity, resulting in effective control of insect pests.IMPORTANCEBacillus thuringiensis took advantage of important insect cellular proteins, such as chaperones, involved in maintaining protein homeostasis, to enhance its insecticidal activity. This constitutes a positive loop where the concentrations of Hsp90 and Hsp70 in the gut lumen are likely to increase as midgut cells burst due to Cry1A pore formation action. Hsp90 protects Cry1A protoxin from degradation and enhances receptor binding, resulting in increased toxicity. The effect of insect chaperones on Cry toxicity could have important biotechnological applications to enhance the toxicity of Cry proteins to insect pests, especially those that show low susceptibility to these toxins.

Draft Genome Sequences and Complete Sequences of Two Plasmids of Escherichia coli Strain BOq 01, a Multiple-Antibiotic-Resistant Strain Isolated from a Poultry Farm in Mexico.

We report here the draft genome sequence of Escherichia coli strain BOq 01, a bacterium isolated from a poultry farm; the genome includes two plasmids conferring antibiotic resistances. This bacterium has a GC content of 50.89% and a genome size of 4.6 Mb.

Draft Genome Sequence of Mycoplasma wenyonii, a Second Hemotropic Mycoplasma Species Identified in Mexican Bovine Cattle.

The hemotropic mycoplasma (hemoplasma) Mycoplasma wenyonii is an animal pathogen that affects bovine cattle health. Here, we present the draft genome sequence of the hemoplasma M. wenyonii strain INIFAP02 found in cattle from Mexico.

Draft Genome Sequences of Endophytic Isolates of Klebsiella variicola and Klebsiella pneumoniae Obtained from the Same Sugarcane Plant.

Endophytic Klebsiella variicola KvMx2 and Klebsiella pneumoniae KpMx1 isolates obtained from the same sugarcane stem were used for whole-genome sequencing. The genomes revealed clear differences in essential genes for plant growth, development, and detoxification, as well as nitrogen fixation, catalases, cellulases, and shared virulence factors described in the K. pneumoniae pathogen.

Draft Genome Sequence of Anaplasma marginale Strain Mex-01-001-01, a Mexican Strain That Causes Bovine Anaplasmosis.

Bovine anaplasmosis is an arthropod-borne hemolytic disease caused by Anaplasma marginale. While only a few Anaplasma marginale strains have been reported, no Mexican strains have been reported. Due to the genetic diversity of A. marginale, the genome of the strain Mex-01-001-01, isolated in Mexico, represents a new source of information.

Draft Genome Sequence of a Pseudomonas aeruginosa NA04 Bacterium Isolated from an Entomopathogenic Nematode.

We report the draft genome sequence of Gram-negative bacterium Pseudomonas aeruginosa NA04, isolated from the entomopathogenic nematode Heterorhabditis indica MOR03. The draft genome consists of 54 contigs, a length of 6.37 Mb, and a G+C content 66.49%.