Fungal community diversity of heavy metal contaminated soils revealed by metagenomics.

The inappropriate disposal of toxic compounds generated by industrial activity has been impacting the environment considerably. Microbial communities inhabiting contaminated sites may represent interesting ecological alternatives for the decontamination of environments. The present work aimed to investigate the fungal diversity and its functionality contained in stream sediments with industrial waste contaminated with heavy metals by using metagenomic approach. A total of 12 fungal orders were retrieved from datasets and, at phylum level, Ascomycota was the most abundant, followed by Basidiomycota, Chytridiomycota and Blastocladiomycota. Higher abundance of sequences was encountered within the less contaminated site, while the lower abundance was found in the sample with the higher contamination with lead. Gene sequences related to DNA repair and heavy metals biosorption processes were found in the four samples analyzed. The genera Aspergillus and Chaetomium, and Saccharomycetales order were highly present within all samples, showing their potential to be used for bioremediation studies. The present work demonstrated the importance of using the metagenomic approach to understand the dynamics and the possible metabolic pathways associated with fungal communities related to environmental samples containing heavy metals, as well as evidenced the importance of improving culturomics techniques for isolating strains with potential application in bioremediation processes of environments contaminated with heavy metals.

First Report of Xanthomonas citri subsp. citri causing Citrus Canker on lime in Rio Grande do Norte, Brazil.

Citrus canker caused by Xanthomonas citri subsp. citri is one of the most important citrus diseases in the world (Gottwald et al. 2002), mainly for citrus-producing countries with humid sub-tropical regions such as United States, Argentina, and Brazil, where losses may be significant (Behlau et al. 2020). In the state of Rio Grande do Norte (RN), Brazil, citrus production is expanding and shows social and economic importance for small farmers, which produced approximately 297 tons of lime in this state in 2019 (IBGE 2021). In December 2019, we observed symptoms of erumpent lesions with margins surrounded by yellow haloes on leaves and fruit of the lime (Citrus aurantifolia cv. 'Galego') (about 5% incidence) in a plantation located in the municipality of Mossoró, RN (05°12'21.1"S, 37°19'16"W). Samples were collected from the lime orchard, and five bacterial strains (CCRMXC01 to CCRMXC05) showing yellow, convex, mucoid colonies were isolated in a nutrient-yeast-dextrose-agar medium (NYDA). Pathogenicity tests were performed on sweet orange (C. sinensis cv. 'Pêra') and lime (C. latifolia cv. 'Tahiti') seedlings. Four wounds per leaf (upper side) were carried out with an entomological pin and 10 µl of a bacterial suspension (108 CFU mL-1) were deposited on each wound. The negative control consisted of leaves treated with sterile distilled water (SDW). For each citrus species, we used four replicates per strain and one leaf with four wounds per replicate. Inoculated leaves developed erumpent lesions with margins surrounded by yellow haloes six days after inoculation (DAI) in both citrus species, while leaves treated with SDW remained symptomless. Nine DAI, we reisolated the pathogen and performed rep-PCR (REP, ERIC, and BOX-PCR) analyses (Gama et al. 2018) with the strains inoculated and reisolated to confirm the identity of the strains and to fulfill Koch's postulates. The strains were stored at the Culture Collection Rosa Mariano (CCRM) of the Phytobacteriology Laboratory at the Universidade Federal Rural de Pernambuco. The five strains reisolated showed the same REP, ERIC, and BOX-PCR profiles as the strains used for inoculations. The molecular identification was performed sequencing the dnaK, fyuA, gyrB, and rpoD genes (Young et al. 2008). Each fragment was sequenced in both the forward and reverse directions. Using the BLASTn tool, we observed that sequences of the dnaK (GenBank MW218913 to MW218917), fyuA (GenBank MW218918 to MW218922), and rpoD (GenBank MW218928 to MW218932) genes of the strains CCRMXC01 to CCRMXC05 showed 100% of identity with the sequences of these genes from the type strain (ICMP 24T) and of other strains of X. citri subsp. citri (ICMP 21 and ICMP 7493), while sequences of gryB (GenBank MW218923 to MW218927) of the former strains showed 100% identity with the gyrB sequence of the strains ICMP 24T and ICMP 7493 and 99,85% identity with strain ICMP 21. This short variation in the sequence of the gyrB gene also may be observed among strains of X. citri subsp. citri available in NCBI database (https://www.ncbi.nlm.nih.gov/). The phylogenetic analysis performed using Bayesian inference and the concatenated sequence of all the type or representative strains of species and pathovars of Xanthomonas available in GenBank showed that the strains CCRMXC01 to CCRMXC05 clustered together with strain ICMP 24T with 1.0 posterior probability. To our information, this is the first report of X. citri subsp. citri causing citrus canker on lime in RN state, Brazil.

Application of next generation sequencing (NGS) for descriptive analysis of 30 genomes of Leishmania infantum isolates in Middle-North Brazil.

Visceral leishmaniasis (VL) is a life-threatening disease caused by the protozoa Leishmania donovani and L. infantum. Likely, L. infantum was introduced in the New World by the Iberic colonizers. Due to recent introduction, the genetic diversity is low. Access to genomic information through the sequencing of Leishmania isolates allows the characterization of populations through the identification and analysis of variations. Population structure information may reveal important data on disease dynamics. Aiming to describe the genetic diversity of L. infantum from the Middle-North, Brazil, next generation sequencing of 30 Leishmania isolates obtained in the city of Teresina, from where the disease dispersed, was performed. The variations were categorized accordingly to the genome region and impact and provided the basis for chromosomal ploidy and population structure analysis. The results showed low diversity between the isolates and the Iberic reference genome JPCM5. Most variations were seen in non-coding regions, with modifying impact. The ploidy number analysis showed aneuploid profile. The population structure analysis revealed the presence of two L. infantum populations identified in Teresina. Further population genetics studies with a larger number of isolates should be performed in order to identify the genetic background associated with virulence and parasite ecology.

Evaluation of the Antibacterial Activity and Efflux Pump Reversal of Thymol and Carvacrol against Staphylococcus aureus and Their Toxicity in Drosophila melanogaster.

The antibacterial activity and efflux pump reversal of thymol and carvacrol were investigated against the Staphylococcus aureus IS-58 strain in this study, as well as their toxicity against Drosophila melanogaster. The minimum inhibitory concentration (MIC) was determined using the broth microdilution method, while efflux pump inhibition was assessed by reduction of the antibiotic and ethidium bromide (EtBr) MICs. D. melanogaster toxicity was tested using the fumigation method. Both thymol and carvacrol presented antibacterial activities with MICs of 72 and 256 µg/mL, respectively. The association between thymol and tetracycline demonstrated synergism, while the association between carvacrol and tetracycline presented antagonism. The compound and EtBr combinations did not differ from controls. Thymol and carvacrol toxicity against D. melanogaster were evidenced with EC50 values of 17.96 and 16.97 µg/mL, respectively, with 48 h of exposure. In conclusion, the compounds presented promising antibacterial activity against the tested strain, although no efficacy was observed in terms of efflux pump inhibition.

Complete Genome Sequence of Xanthomonas citri pv. anacardii Strain IBSBF2579 from Brazil.

The bacterium Xanthomonas citri pv. anacardii is the agent of angular leaf spot of the cashew tree (Anacardium occidentale L.). The complete genome sequencing of the strain IBSBF2579 was done on an Illumina HiSeq 2500 platform. The de novo assembly of the X. citri pv. anacardii strain IBSBF2579 genome yielded 133 contigs, with a size of 5,329,247 bp and a G+C content of 64.03%. The prediction was performed by GeneMarkS and the automatic annotation by Rapid Annotations using Subsystems Technology (RAST), with 4,406 identified genes.

Novel E6 and E7 oncogenes variants of human papillomavirus type 31 in Brazilian women with abnormal cervical cytology.

HPV-31 has been widely described as an important oncogenic type, showing high incidence in worldwide and especially in Northeastern Brazil. We sought to identify the presence of specific mutations in HPV-31 E6 and E7 oncogenes in women with abnormal cervical smear. We enrolled 150 gynecological patients from Sergipe State, Northeastern Brazil. HPV screening was carried out by polymerase chain reaction (MY09/11). E6 and E7 oncogenes were amplified with specific primers and sequenced. The sequences obtained were aligned with the GenBank reference sequences in order to search for genetic variants. We identified genetic variants in E6 and E7 sequences from HPV-31. Two new nucleotide changes in E6 and E7 were described for the first time in this study. A novel mutation in E6 resulted in amino acid change in a site belonging to T-cell epitope with MHC II binding activity. There was no significant difference in the distribution of HPV-31 E6 and E7 variants when compared to all selected clinical/epidemiological characteristics. HPV-31 isolates have been clustered into three main groups called lineages A, B and C. We describe new HPV-31 variants in Brazil, contributing to better understand the genomic diversity of these viruses.

A list of mosquito species of the Brazilian State of Pernambuco, including the first report of Haemagogus janthinomys (Diptera: Culicidae), yellow fever vector and 14 other species (Diptera: Culicidae).

INTRODUCTION: Besides mosquito species adapted to urban environments (Culex quinquefasciatus, Aedes aegypti and Aedes albopictus), only 15 species of Anopheles had been recorded in the State of Pernambuco. METHODS: Human-landing mosquitoes were collected in Dois Irmãos Park, in Recife. RESULTS: The first report for the state of Haemagogus janthinomys, an important vector of yellow fever virus, and 14 other species, including Trichoprosopon lampropus, a first reported for Brazil. CONCLUSIONS: The mosquito fauna in the area is diversified and has potential medical and veterinary importance.

A list of mosquito species of the Brazilian State of Pernambuco, including the first report of Haemagogus janthinomys (Diptera: Culicidae), yellow fever vector and 14 other species (Diptera: Culicidae) / Lista de espécies de mosquitos do Estado de Pernambuco e primeiro relato de Haemagogus janthinomys (Diptera: Culicidae) vetor de febre amarela silvestre e outras 14 espécies (Diptera: Culicidae)

INTRODUCTION: Besides mosquito species adapted to urban environments (Culex quinquefasciatus, Aedes aegypti and Aedes albopictus), only 15 species of Anopheles had been recorded in the State of Pernambuco. METHODS: Human-landing mosquitoes were collected in Dois Irmãos Park, in Recife. RESULTS: The first report for the state of Haemagogus janthinomys, an important vector of yellow fever virus, and 14 other species, including Trichoprosopon lampropus, a first reported for Brazil. CONCLUSIONS: The mosquito fauna in the area is diversified and has potential medical and veterinary importance.

INTRODUÇÃO: Além de mosquitos adaptados ao ambiente urbano (Culex quinquefasciatus, Aedes aegypti e Ae. albopictus), apenas 15 espécies de Anopheles haviam sido relatadas no Estado de Pernambuco. MÉTODOS: Mosquitos que pousavam em humanos no Parque Dois Irmãos, em Recife foram coletados. RESULTADOS: Haemagogus janthinomys, importante vetor de vírus de febre amarela, e outras 14 espécies são relatadas pela primeira vez no estado, incluindo Trichoprosopon lampropus, relatado pela primeira vez no Brasil. CONCLUSÕES: A fauna de mosquitos na área é muito diversificada e tem potencial importância médica e veterinária.

Description of three practical and inexpensive devices for the collection of mosquitoes and other small insects.

Three new devices for collecting and maintaining mosquitoes and other small insects are described. A suction tube, composed of two pieces of plastic transparent tube, the narrower fitted in the wider, with the mouth protected by the use of voile, is cheap and practical and can be transported around the neck. A 900-ml plastic transparent box is utilized for the restraint of sucked insects and, with the adaptation of two pieces of plastic tube, as a suction chamber.