Polymorphic DNA Analysis Research Articles

Biodiversity among Brettanomyces bruxellensis Strains Isolated from Different Wine Regions of Chile: Key Factors Revealed about its Tolerance to Sulphite.

Brettanomyces bruxellensis is regarded as the main spoilage microorganism in the wine industry, owing to its production of off-flavours. It is difficult to eradicate owing to its high tolerance of adverse environmental conditions, such as low nutrient availability, low pH, and high levels of ethanol and SO2. In this study, the production of volatile phenols and the growth kinetics of isolates from various regions of Chile were evaluated under stressful conditions. Through randomly amplified polymorphic DNA (RAPD) analysis, 15 strains were identified. These were grown in the presence of p-coumaric acid, a natural antimicrobial and the main precursor of off-flavours, and molecular sulfur dioxide (mSO2), an antimicrobial synthetic used in the wine industry. When both compounds were used simultaneously, there were clear signs of an improvement in the fitness of most of the isolates, which showed an antagonistic interaction in which p-coumaric acid mitigates the effects of SO2. Fourteen strains were able to produce 4-vinylphenol, which showed signs of phenylacrylic acid decarboxylase activity, and most of them produced 4-ethylphenol as a result of active vinylphenol reductase. These results demonstrate for the first time the serious implications of using p-coumaric acid, not only for the production of off-flavours, but also for its protective action against the toxic effects of SO2.

Genetic diversity and toxic genes analysis of vibrio spp. isolated from white leg shrimp and marine fishes cultured in Tam G iang lagoon in Thua Thien Hue province, Vietnam

Objective: This study was done to report the results of genetic diversity and toxic genes analysis of Vibrio pathogen isolated from white leg shrimp and marine fishes cultured in Thua Thien Hue province, Vietnam. Methods/statistical analysis: Pathogen Vibrio spp. were isolated from shrimps and fishes, and were identified by 16S rRNA sequencing. The presence of toxin genes in Vibrio spp. strains were determined through the presence of genes encoding toxic proteins (pirAvp, pir-Bvp, tlh, tdh and trh) based on specific primers for these genes. Genetic diversity of Vibrio strains was analysed by RAPD technique. Findings: A total of 120 Vibrio colonies from shrimps (with Acute Hepatopancreatic Necrosis Disease) and fishes (with hemorrhagic disease) cultured in Tam Giang lagoon in Thua Thien Hue, Vietnam were isolated. Of which, 14/54 strains from shrimps had pirAvp and pirBvp genes and 18/66 strains from fishes had tlh gene, and none of Vibrio strains had tdh and trh genes. Randomly amplified polymorphic DNA (RAPD) analysis of 36 Vibrio samples showed the 148 polymorphic DNA fragments from ten random primers. The genetic diversity is high within studied species. In there, V. parahaemolyticus has the highest diversity level (h=0.1645 and I=0.2695) while V. shilonii is lowest (h=0.0136) and I=0.0207). The degree of genetic differentiation among species is also high (Gst=0.4827). Genetic identity between V. parahaemolyticus and V. vulnificus (0.9545) is highest while between V. shilonii and V. harveyi (0.7416) is lowest. The dendrogram also showed that V. parahaemolyticus is closely related to V. vulnificus whereas V. shilonii and V. harveyi have large distance. Application/improvements: This study is aimed to provide scientific data as the basis for the study and production of rapid diagnostic kits in the future. Keywords: AHPND; Genetic diversity; RAPD; toxic genes; Vibrio; Vietnam

Estimation of Morphological and Molecular Variability in induced Mutant Fennel Lines

In the present study, M3 mutant lines generated from gamma radiation of widely adopted fennel cultivar Rajasthan Fennel-125 (RF- 125) was used. High variability was recorded for time of flowering, number of primary and secondary branches, number of umbels and umbellate/plant, number of seeds/umbel, plant height, days to maturity and seed yield/plant. Total of 24 diverse mutants were further selected for molecular characterization based on morphological diversity. The Random Amplified Polymorphic DNA (RAPD) analysis showed broad genetic diversity among the mutant lines. Total 73 loci were amplified in all 24 mutants and 23 loci were found polymorphic. Dendrogram showed similarity coefficient value in range of 0.00 to 0.88. The dendrogram divided the 24 mutants into four main clusters. The morphological and molecular study revealed that the mutants selected in the M2 generation showed variability at the genetic level for different essential traits of fennel.

Evolutionary perspectives on bee mtDNA from mito-OMICS analyses of a solitary species

The analysis of mitochondrial DNA polymorphism has been applied in multiple organisms to obtain information about species biology, ecology, population dynamics, and evolution. In this manuscript, the complete sequencing and characterization of the mitochondrial genome (mtGenome) of Tetrapedia diversipes are reported and discussed from comparative and evolutionary perspectives among all mtGenomes available for bees so far. The T. diversipes mtGenome is 15,358 bp long and exhibits the typical set of genes and an A+T-rich region of 588 bp. The overall base composition is biased towards A/T (84.3%), with 42.6% A, 41.7% T, 9.8% C, and 5.9% G nucleotides. The obtained data also comprise the mitochondrial DNA methylation and single-nucleotide polymorphic sites of one T. diversipes population. Transcription follows the “tRNA punctuation” model, with at least three primary polycistronic transcripts that are posteriorly processed. Additionally, higher expression rates of the 16S gene suggest the existence of an exclusive transcription site in this region, and the differential expression of the 12S gene between larvae and adults reveals different isoforms for this gene. The sequence order of protein-coding and rRNA genes is conserved across different bee lineages, and differences are restricted to tRNA gene positions. The present results characterize numerous understudied aspects of bee mtGenomes, and a major evolutionary review of this molecule within the group is provided. Therefore, this work is a valuable resource for studying mitochondrial molecular biology and evolution in bees.

Corneal ulcer/keratitis derived Aspergillus flavus &Aspergillus tamarii and their RAPD-PCR typing

The aim of the study was investigate the DNA polymorphisms of Aspergillus flavus and Aspergillus tamarii isolated from regional fungal keratitis cases based on random amplified polymorphic DNA (RAPD) analyses. Thirty-two A. flavus and six A. tamarii isolates were identified using conventional macroscopic and microscopic features. Five decamer primers were used to amplify the genomic DNA sequences of the test isolates, and 140 total RAPD patterns were generated among them. From each of the 38 isolates, a range of 15–30 RAPD types were revealed. All of these were isolated from male farmers from Tamil Nadu. In the future, RAPD analyses could be employed as a reliable and rapid technique to differentiate among keratitis isolates of fungal pathogens.

Changes in essential oil composition, phenylalanine ammonia lyase gene expression and rosmarinic acid content during shoot organogenesis in cytokinin-treated Satureja spicigera (C. Koch) boiss. shoots

This study investigated the effects of different concentrations of zeatin and thidiazuron on the accumulation of phenolics and activity of phenylalanine ammonia lyase gene in the micropropagated Satureja spicigera shoots. S. spicigera clones were generated from cultured nodal segments on Murashige and Skoog including vitamins medium supported with zeatin and thidiazuron concentrations. Shoots were subcultured three times in the same medium and harvested at the end of the third month. The expression levels of phenylalanine ammonia lyase gene were conducted using semi-quantitative reverse transcription polymerase chain reaction technique. Rosmarinic acid contents were determined using reverse phase-high performance liquid chromatography with a diode array detector. Genetic stability of regenerants was performed using random amplified polymorphic DNA markers. Essential oils from shoots were identified by gas chromatography–mass spectrophotometry analysis. It has been detected that 0.1 mg/L thidiazuron is more effective on the node formation and shoot elongation. Contrarily, 4.0 mg/L thidiazuron caused a significant increase in shoot multiplication. The highest biomass accumulation was calculated from medium including 1.0 mg/L zeatin. According to random amplified polymorphic DNA analysis, all banding profiles from micropropagated plants were monomorphic. Gas chromatography–mass spectrophotometry analysis showed that rising concentrations of zeatin and thidiazuron increased the thymol and decreased the carvacrol content. It was also found that rosmarinic acid production increased significantly at 1.0 mg/L zeatin application. However, thidiazuron applications had no positive effect on rosmarinic acid accumulation. Comparing phenylalanine ammonia lyase gene expression and rosmarinic acid accumulation in in vitro shoots, it was found that they were correlated.

Assessment of waste frying oil transesterification capacities of local isolated Aspergilli species and mutants

Abstract Biodiesel (fatty acids methyl esters, FAME) has attracted considerable attention as an environmentally and eco-friendly alternative for diesel engines. This research manipulates the use of Aspergillus whole-cell lipase as a biocatalyst for biodiesel production from waste frying oil (WFO). A total of 17 isolates of Aspergillus species screened for lipase and esterase production abilities. Qualitatively, 11 Aspergillus isolates showed lipase and/or esterase activities and only 4 isolates were able to perform WFO transesterification under the tested conditions. Two Aspergillus isolates showed relatively high FAME yields, thus were selected as good enzyme producers. These two isolates were molecularly identified using rRNA gene sequence ITS1 and ITS2 as A. tamarii NDA03a (Genbank Accession Number MK849615) and A. flavus NDA04a (Genbank Accession Number MK811208), respectively. These identified isolates were exposed to ethyl methanesulfonate (EMS) for producing hyper lipolysis mutants. Mutagenesis led to 13.15 and 14.45% improvement of the WFO transesterification by A. tamarii NDA03a and A. flavus NDA04a, respectively. Random amplified polymorphic DNA (RAPD) analysis of the produced mutants confirmed the genetic basis of the activity variation. Genetic polymorphism reached to 79.31% and 80.65% between A. flavus NDA04a and A. tamarii NDA03a mutants and their corresponding wild types, respectively.

Chemical mutagenesis of Saccharomyces cerevisiae for enhancing bioethanol production with fermentation at very high sugar concentration

Saccharomyces cerevisiae is one of the most promising unicellular fungi on account of its vital applications in biotechnology as well as bioethanol production. Improvement of ethanol production via very high-gravity (VHG) fermentation (fermentation at high sugar levels) was successfully developed using the ethidium bromide (EtB) mutagenesis of S. cerevisiae. This study found two developed mutants of S. cerevisiae (EtB20a and EtB20b) with varied capacity for ethanol production using EtB, depending on random amplified polymorphic DNA analysis. Mutant EtB20b showed improved ethanol yield (19.5%) compared with the wild-type (18.0%), while the other mutant EtB20a exhibited retarded ethanol production (9.1%). Optimization of ethanol production by mutant EtB20b was performed under other conditions including temperature, pH, inoculum size, and incubation period. The highest production capacity of the yeasts was 20.8, 19.9, 19.5, and 19.5% at an optimum temperature of 30 °C, pH 6.0, incubation period of 72 h, and 1 mL of yeast suspension (optical density at 600 nm) with glucose utilization of 42.6, 40.7, 39.8, and 39.9%, respectively.

Rapid identification of the indigenous medicinal crop Cinnamomum osmophloeum from various adulterant Cinnamomum species by DNA polymorphism analysis

Background: Cinnamomum osmophloeum (Co), a member of the Lauraceae, is an indigenous medicinal crop in Taiwan, and it contains higher cinnamaldehyde in essential oil than do other Cinnamomum species. Among these species, Cinnamomum burmannii (Cb) is frequently adulterated as Co because of their similar morphological characteristics or features. Objective: The purpose of this study was to develop a DNA-based molecular method for rapid authentication of the indigenous Co and prevention of its adulteration. Materials and Methods: The internal transcribed spacer (ITS) regions of nuclear ribosomal DNA from various Cinnamomum species were amplified by polymerase chain reaction (PCR), and these obtained sequences were used for sequence analysis. Based on the sequence variants among various Cinnamomum species, restriction fragment length polymorphism (RFLP) was used to differentiate these Cinnamomum plants. Results: Two restriction endonucleases, MylI and EcoRV, were specifically used to digest the PCR-amplified ITS DNA from seven Cinnamomum species. The PCR-RFLP results demonstrated that the different restriction patterns that were produced by these two restriction enzymes clearly distinguished Co from Cb and five other Cinnamomum species simultaneously. Conclusion: The PCR-RFLP analysis developed in this study provides an alternative method for rapidly identifying Cinnamomum plants at the species level using DNA polymorphisms.

Bio-nanoparticle assembly: a potent on-site biolarvicidal agent against mosquito vectors.

Background: Vector-borne diseases such as malaria, dengue, yellow fever, encephalitis and filariasis are considered serious human health concerns in the field of medical entomology. Controlling the population of mosquito vectors is one of the best strategies for combating such vector-borne diseases. However, the use of synthetic insecticides for longer periods of time increases mosquito resistance to the insecticides. Recently, the search for new environmentally friendly and efficient insecticides has attracted major attention globally. With the evolution of material sciences, researchers have reported the effective control of such diseases using various sustainable resources. The present investigation demonstrates a potent on-site biolarvicidal agent against different mosquito vectors such as Aedes albopictus, Anopheles stephensi and Culex quinquefasciatus. Methods: Stable and photo-induced colloidal silver nanoparticles were generated via the surface functionalization of the root extract of Cyprus rotundas. Characterizations of the nanoparticles were performed using assorted techniques, such as UV-visible spectroscopy, FTIR spectroscopy, DLS and HRTEM. The bioefficacy of the synthesized nanoparticles was investigated against different species of mosquito larvae through the evaluation of their life history trait studies, fecundity and hatchability rate of the treated larvae. Histopathological and polymerase chain reaction-random amplified polymorphic DNA (RAPD) analyses of the treated larvae were also examined to establish the cellular damage. Results: The synthesized nanoparticles showed remarkable larvicidal activity against mosquito larvae in a very low concentration range (0.001–1.00) mg L−1. The histopathological study confirmed that the present nanoparticles could easily enter the cuticle membrane of mosquito larvae and subsequently obliterate their complete intestinal system. Furthermore, RAPD analysis of the treated larvae could assess the damage of the DNA banding pattern. Conclusion: The present work demonstrates a potent biolarvicidal agent using sustainable bioresources of the aqueous Cyprus rotundas root extract. The results showed that the synthesized nanoparticles were stable under different physiological conditions such as temperature and photo-induced oxidation. The effectiveness of these materials against mosquito larvae was quantified at very low dose concentrations. The present biolarvicidal agent can be considered as an environmentally benign material to control the mosquito vectors with an immense potential for on-site field applications.

Short tandem repeat typing of cells transferred via micromanipulation with whole-genome amplification.

This study attempted to determine the minimum number of cells required to conduct DNA analyses effectively. Oral mucosal cells obtained from eight persons were suspended and individually collected by using micromanipulation technique. DNA was extracted and amplified by whole-genome amplification (WGA). Nuclear DNA was extracted to evaluate the feasibility of autosomal short tandem repeat (STR) polymorphism and Y-chromosomal STR polymorphism analyses. Tests were conducted with 20 and 30 cells, to determine the minimum number of cells required for each DNA analysis. Tests with 20 cells were repeated 5 times, to examine reproducibility. When five or 10 cells were used, loci could not be identified for most alleles. Furthermore, DNA polymorphism analyses of a single cell transferred directly to a polymerase chain reaction solution were unsuccessful. The present findings suggest that, in forensic identification, 20 or more cells are required in order to obtain clear results from autosomal and Y-chromosomal STR polymorphism analyses. Furthermore, the feasibility of sample preservation and reexamination was also confirmed by DNA amplification with WGA.

Diversity of nitrogen-fixing bacteria in agricultural field soil along the Yamuna River, Delhi, India

Nitrogen-fixing bacteria play an important role in sustaining soil health. The diversity was studied in nitrogen-fixing bacteria using the nifH gene along the Yamuna River in Delhi-National Capital Region. The nifH gene is a part of the nif regulon, which codes for an Fe-protein of the nitrogenase enzyme complex, responsible for the reduction of dinitrogen (N2) to ammonia (NH3) during biological nitrogen fixation. Soil samples were collected from agricultural fields near the Yamuna River. Terminal restriction fragment length polymorphism analysis of soil DNA using the TReFID software package showed community comprising different nitrogen-fixing bacterial phyla, dominated by the Proteobacteria (26.87%) and Actinobacteria (2.9%). The relative abundance of these two phyla increased downstream along the river while the relative abundance of the Cyanobacteria decreased. The maximum overall microbial diversity index was lower downstream and during the summer season. It was concluded that the seasonal variation in temperature and moisture and the increases in the pollution of the water directly affected the microbial diversity as well as the relative abundance of the nitrogen-fixing bacterial population in agricultural fields along the Yamuna River.

Co-Isolation and Characterization of Two Pandoraviruses and a Mimivirus from a Riverbank in Japan

Giant viruses, like pandoraviruses and mimiviruses, have been discovered from diverse environments, and their broad global distribution has been established. Here, we report two new isolates of Pandoravirus spp. and one Mimivirus sp., named Pandoravirus hades, Pandoravirus persephone, and Mimivirus sp. isolate styx, co-isolated from riverbank soil in Japan. We obtained nearly complete sequences of the family B DNA polymerase gene (polB) of P. hades and P. persephone; the former carried two known intein regions, while the latter had only one. Phylogenetic analysis revealed that the two new pandoravirus isolates are closely related to Pandoravirus dulcis. Furthermore, random amplified polymorphic DNA analysis revealed that P. hades and P. persephone might harbor different genome structures. Based on phylogenetic analysis of the partial polB sequence, Mimivirus sp. isolate styx belongs to mimivirus lineage A. DNA staining suggested that the Pandoravirus spp. asynchronously replicates in amoeba cells while Mimivirus sp. replicates synchronously. We also observed that P. persephone- or Mimivirus sp. isolate styx-infected amoeba cytoplasm is extruded by the cells. To the best of our knowledge, we are the first to report the isolation of pandoraviruses in Asia. In addition, our results emphasize the importance of virus isolation from soil to reveal the ecology of giant viruses.

Conventional and qPCR reveals the presence of ‘Candidatus Liberibacter solanacearum’ haplotypes A, and B in Physalis philadelphica plant, seed, and Βactericera cockerelli psyllids, with the assignment of a new haplotype H in Convolvulaceae

The husk tomato (Physalis philadelphica Lam.) is an important Solanaceae native to Mesoamerica that is grown for its green fruit used as an important ingredient in domestic and international cuisine. Nevertheless, husk tomato plants with symptoms resembling those caused by 'Candidatus Liberibacter solanacearum' (CLso) have been observed during the last decade in plantations located in the State of Mexico, Michoacan and Sinaloa in Mexico. These areas are located near other solanaceous crops where Bactericera cockerelli the well-known psyllid transmitter of CLso is frequently present. Thus, the goal of this study was to determine if CLso haplotypes are present in husk tomato varieties in commercial fields in Mexico. From 2015 to 2016, plants and fruit showing evident symptoms of CLso infection, as well as psyllids were collected in these states and assayed by PCR for CLso using primer sets OA2/OI2c and LpFrag 1-25F/427R. Phylogenetic reconstruction was performed with Bayesian analysis and maximum likelihood methods using amplicon sequences obtained in this work along with those deposited in the GenBank database corresponding to the CLso detected in Solanaceae, Apiaceae, and Convolvulaceae host families. In addition, all the sequences were subjected to haplotype determination through an analysis of DNA polymorphisms using the DnaSP software. Furthermore, quantitative PCR (qPCR) was performed using CLso-specific primers and probes. Phylogenetic reconstruction and qPCR confirmed the presence of CLso in plants, seeds and insect-vectors, and CLso sequences from plants and seeds completely matched haplotype B, whereas CLso haplotypes A and B were detected in B. cockerelli psyllids. Polymorphism analysis identified a novel Convolvulaceae-associated CLso haplotype, which was named haplotype H. The results of this study will enable the dissemination of infected seeds to new husk tomato production areas to be avoided.

Genetic polymorphisms in mutagenesis progeny of Arabidopsis thaliana irradiated by carbon-ion beams and γ-rays irradiations

Purpose: Heavy-ion beams and γ-rays are popular physical mutagenesis to generate mutations in higher plants. It has been found that they show different mutation frequencies and spectrums of phenotype induction, however, the characteristics of heavy-ion beams on genetic polymorphism have not been clarified by comparing with γ-rays.Materials and methods: In the present study, seeds of Arabidopsis thaliana were exposed to carbon-ion beams (with linear energy transfer (LET) of 50 keV/μm) and γ-rays (with average LET of 0.2 keV/μm) irradiation. By using inter-simple sequence repeat (ISSR) and random amplified polymorphic DNA (RAPD) analysis, the genetic polymorphism of both M1 and M3 plants were investigated, respectively.Results: Carbon-ion beams induced relatively higher polymorphism rate in both M1 and M3 generation than γ-rays: the polymorphism rates of M1 plants derived from carbon-ion beams irradiation are 12.87% (ISSR-C) and 9.01% (RAPD-C), while are 7.67% (ISSR-γ) and 1.45% (RAPD-γ) of plants derived from γ-rays. In M3 generation, the polymorphism rates of ISSR-C, RAPD-C, ISSR-γ, and RAPD-γ are 17.64%, 22.79%, 12.10%, and 2.82%, respectively.Conclusions: In summary, the exposure to carbon-ion beams and γ-rays lead to the change of genomic DNA of A. thaliana, which could be tested in M1 plants and M3 plants by ISSR and RAPD technology. So, both carbon-ion beams and γ-rays can induce variations of genetic polymorphisms in M1 plants and M3 plants. The genetic polymorphisms of M1 plants and M3 plants induced by carbon-ion beams are higher than γ-rays, indicating that heavy-ion beams irradiations mutation breeding is more advantageous than conventional ionizing radiations. Average molecular polymorphism of M1 plants is lower than M3 mutants, by nearly 4.77% (ISSR-C), 13.78% (RAPD-C), 4.43% (ISSR-γ), and 1.37% (RAPD-γ). We hope our study will provide basic information for understanding the effects of carbon-ion beams and γ-rays for plant mutation breeding.

MUS81 Participates in the Progression of Serous Ovarian Cancer Associated With Dysfunctional DNA Repair System.

Objective: Methyl methanesulfonate ultraviolet sensitive gene clone 81 (MUS81) is a structure-specific endonuclease that plays a pivotal role in the DNA repair system of cancer cells. In this study, we aim to elucidate the potential association between the dysfunction of MUS81 and the progression of Serous Ovarian Cancer (SOC).Methods: To investigate the association between MUS81 and prognosis of SOC, immunohistochemistry technology and qPCR were used to analyze the level of MUS81 expression, and transcriptional profile analysis and protein interaction screening chip were used to explore the MUS81 related signal pathways. Random amplified polymorphic DNA (RAPD) analysis, immunofluorescence and comet assays were further performed to evaluate genomic instability and DNA damage status of transduced SOC cells. Experiments both in vitro and in vivo were conducted to verify the impact of MUS81 silencing on chemotherapeutic drug sensitivity of SOC.Results: The overexpression of MUS81 in SOC tissues was related to poor clinical outcomes. The transcriptional chip data showed that MUS81 was involved in multiple pathways associated with DNA repair. Deficiency of MUS81 intensified the genome instability of SOC cells, promoted the emergence of DSBs and restrained the formation of RAD51 foci in SOC cells with exposure to UV. Furthermore, downregulation of MUS81 enhanced the sensitivity to Camptothecin and Olaparib in SOC cell lines and xenograft model.Conclusions: MUS81 is involved in the progression of SOC and inhibition of MUS81 could augment the susceptibility to chemotherapeutic agents. MUS81 might represent a novel molecular target for SOC chemotherapy.

Is Genomic DNA extracted and stored at -20°C for long time useful in forensic field?

With the progress of technologies, forensic genetic laboratories have been more frequently involved in "cold case". The aim of this study was to understand whether the DNA extracted and stored for a long time at−20 °C could be useful for new analyses in order to identify the perpetrator of unsolved crimes, especially when evidence is no longer available. We selected 120 DNA samples obtained from evidence collected at crime scene between 2001 and 2010: they were quantified again using Realtime PCR and the “Plexor HY System” kit. The results were compared with those obtained in the past. For 37 samples discrepancies were observed (i.e. positive quantification, identification of male material mixed with female ones). Then these samples were amplified with “AmpFISTR Identifiler Plus” and “PowerPlex ESI 17 Fast System” kits. Genetic profiles useful for a comparison were obtained for all the 37 analysed samples. In 5 samples mixed profiles were highlighted, unlike what was obtained in the past, and in addition, in 28 cases, where no genetic profile was previously typed, a genetic profile was also identified. The study shows how it is possible, even after a considerable time interval, to still obtain genetic profiles useful for a comparison, as well as the possibility of typing new ones. Comparing the results of the analysis of DNA polymorphisms it is possible to underline the improvement that the current techniques have brought to the typing of genetic profiles: regarding the attribution of gender and the perception of contributors within mixtures.

Current Status of Soybean Anthracnose Associated with Colletotrichum truncatum in Brazil and Argentina.

Brazil and Argentina have a combined soybean area of 53.6 million hectares, which accounts for over half of the total global production. The soybean crop in South America extends from latitude 8–10° S to 32–36° S. Such a vast, almost contiguous area imposes a serious sanitary risk to the crop. Currently, the prevalence of anthracnose is increasing, with recurring reports of severe epidemics and expressive yield losses. Soybean anthracnose is mainly associated with Colletotrichum truncatum, although other Colletotrichum species have also been reported as causal agents of this disease. Knowledge about the morphological, cultural, and molecular variability of C. truncatum in South America is crucial for disease management. Here, we present data on the molecular, morphological, biological, cultural, and pathogenicity of C. truncatum isolates collected in Brazil and Argentina. Light microscopy and randomly-amplified polymorphic DNA (RAPD) analysis were used for estimating the variability of isolates. Colletotrichum truncatum displayed three types of conidiogenesis, viz. conidial formation from conidiogenous cells on hyphal extremities, in conidiomas in acervuli, and directly from fertile setae (a mechanism yet-unreported for C. truncatum). RAPD profiling was effective in revealing the genetic diversity among C. truncatum isolates. The intra-group similarity was greater among the Argentinian isolates when compared to the Brazilian group. Furthermore, the results indicated a strong correlation between geographical origin and molecular grouping, with the exclusive or semi-exclusive assembling of Brazilian and Argentinian isolates in distinct clades. Finally, a preliminary account of the reaction of soybean accessions to C. truncatum is also included.

2465. Healthcare-Associated Pediatric Cutaneous Mucormycosis at Texas Children’s Hospital, 2012–2019

BackgroundCutaneous mucormycosis in children is a rare fungal infection which primarily occurs in patients with underlying medical conditions and is associated with significant morbidity. We describe characteristics of pediatric patients with healthcare-associated (HCA) cutaneous mucormycosis at Texas Children’s Hospital (TCH) and results of an outbreak investigation.MethodsPatients at TCH were identified retrospectively through review of the TCH Microbiology Laboratory mycology culture reports from 1/1/2012–2/18/2019. Pediatric patients <21 years of age with cutaneous mucormycosis that developed during a hospitalization or was associated with a medical device were included. Demographic information was collected through review of the electronic medical record. Randomly amplified polymorphic DNA (RAPD) analysis were performed by the Fungus Testing Laboratory, University of Texas Health Science Center at San Antonio. This study was approved through the Baylor College of Medicine Institutional Review Board.ResultsWe identified 12 patients with HCA cutaneous mucormycosis. The characteristics of these patients are detailed in the table. Six cases of Rhizopus infection occurred over a narrow 7 month period from March through September of 2017 prompting an outbreak investigation. Genotyping results revealed 4 molecular types from 5 available isolates suggesting that the isolates were not clonally related. Adhesive products were suspected as a potential source. Infection Control activities included site visits to the hospital supply warehouse and linen facility. Tape practices throughout the hospital were assessed through a nursing survey. Samples of adhesive products were cultured at an environmental microbiology laboratory and no Rhizopus spp. were isolated. A source was not identified.ConclusionMucormycosis is a life-threatening infection in children. Providers should have a low threshold of suspicion for cutaneous mucormycosis in patients with underlying medical conditions (malignancies and extreme prematurity) that develop skin lesions near medical device dressings or securement sites. Disclosures All authors: No reported disclosures.

Genotyping and antimicrobial resistance of Streptococcus uberis isolated from bovine clinical mastitis.

A genotypic characterization of Streptococcus uberis isolated from clinical mastitis (CM) in dairy cows, and the association of Strep. uberis genotypes and antimicrobial susceptibility (AMS) was performed. A total of 89 isolates identified as Strep. uberis from 86 dairy cows with CM in 17 dairy herds of Southeastern Brazil were genotyped using random amplified polymorphic DNA (RAPD) analysis. After genotyping, two clusters (I and II) were created according to RAPD types. A commercial broth microdilution test was used to determine the susceptibility of Strep. uberis isolates to 8 antimicrobials (ampicillin, ceftiofur, cephalothin, erythromycin, penicillin, penicillin+novobiocin, pirlimycin and tetracycline). For each antimicrobial, we determined the minimal inhibitory concentrations that inhibit 50% (MIC50) and 90% (MIC90) of Strep. uberis strains. Differences in AMS among genotypic clusters were evaluated using mixed regression models. Overall, a great polymorphism (56 RAPD-types) was found among Strep. uberis isolates, although a higher genetic similarity (based on the PCR bands features) was observed within herds after genotypic clustering. No differences in AMS were observed among clusters. Strep. uberis isolated from bovine CM were resistant to most antimicrobials, with the exception of cephalothin and penicillin+novobiocin.