Omics analysis coupled with gene editing revealed potential transporters and regulators related to levoglucosan metabolism efficiency of the engineered Escherichia coli

Background Bioconversion of levoglucosan, a promising sugar derived from the pyrolysis of lignocellulose, into biofuels and chemicals can reduce our dependence on fossil-based raw materials. However, this bioconversion process in microbial strains is challenging due to the lack of catalytic enzyme relevant to levoglucosan metabolism, narrow production ranges of the native strains, poor cellular transport rate of levoglucosan, and inhibition of levoglucosan metabolism by other sugars co-existing in the lignocellulose pyrolysate. The heterologous expression of eukaryotic levoglucosan kinase gene in suitable microbial hosts like Escherichia coli could overcome the first two challenges to some extent; however, no research has been dedicated to resolving the last two issues till now. Results Aiming to resolve the two unsolved problems, we revealed that seven ABC transporters (XylF, MalE, UgpB, UgpC, YtfQ, YphF, and MglA), three MFS transporters (KgtP, GntT, and ActP), and seven regulatory proteins (GalS, MhpR, YkgD, Rsd, Ybl162, MalM, and IraP) in the previously engineered levoglucosan-utilizing and ethanol-producing E. coli LGE2 were induced upon exposure to levoglucosan using comparative proteomics technique, indicating these transporters and regulators were involved in the transport and metabolic regulation of levoglucosan. The proteomics results were further verified by transcriptional analysis of 16 randomly selected genes. Subsequent gene knockout and complementation tests revealed that ABC transporter XylF was likely to be a levoglucosan transporter. Molecular docking showed that levoglucosan can bind to the active pocket of XylF by seven H-bonds with relatively strong strength. Conclusion This study focusing on the omics discrepancies between the utilization of levoglucosan and non-levoglucosan sugar, could provide better understanding of levoglucosan transport and metabolism mechanisms by identifying the transporters and regulators related to the uptake and regulation of levoglucosan metabolism. The protein database generated from this study could be used for further screening and characterization of the transporter(s) and regulator(s) for downstream enzymatic/genetic engineering work, thereby facilitating more efficient microbial utilization of levoglucosan for biofuels and chemicals production in future.

Surveillance, Epidemiology and Impact of EV-A71 Vaccination on Hand, Foot, and Mouth Disease in Nanchang, China, 2010–2019

After the first national-scale outbreak of Hand, foot, and mouth disease (HFMD) in China, a national surveillance network was established. Here we described the epidemiology and pathogenic profile of HFMD and the impact of EV-A71 vaccination on pathogen spectrum of enteroviruses in the southeastern Chinese city of Nanchang during 2010–2019. A total of 7,951 HFMD cases from sentinel hospitals were included, of which 4,800 EV-positive cases (60.4%) were identified by real-time RT-PCR. During 2010–2012, enterovirus 71 (EV-A71) was the main causative agent of HFMD, causing 63.1% of cases, followed by 19.3% cases associated with coxsackievirus A16 (CV-A16). Since 2013, the proportion of other enteroviruses has increased dramatically, with the sub genotype D3 strain of Coxsackievirus A6 (CV-A6) replacing the dominance of EV-A71. These genetically diverse native strains of CV-A6 have co-transmitted and co-evolved in Nanchang. Unlike EV-A71 and CV-A16, most CV-A6 infections were concentrated in autumn and winter. The incidence of EV-A71 infection negatively correlated with EV-A71 vaccination (r = −0.990, p = 0.01). And severe cases sharply declined as the promotion of EV-A71 vaccines. After 2-year implementation of EV-A71 vaccination, EV-A71 is no longer detected from the reported HFMD cases in Nanchang. In conclusion, EV-A71 vaccination changed the pattern of HFMD epidemic, and CV-A6 replaced the dominance of EV-A71 over time.

Morphological variation of Aceh Pinus (Pinus merkusii)

The genus Pinus is classified as one of the most widely distributed genera of conifer trees, that are majorly distributed in the Northern and Southern hemispheres. Pinus merkusii is the only member of the genus Pinus that is adapted in the tropics and grows naturally in south Equator. Its natural distribution found through-out Southeast Asia, e.g. in Indonesia; particularly on Sumatran Island. Three native strains are known on Sumatra: Aceh, Kerinci, and Tapanuli; with the first one is known as the largest population among the three and grows naturally on the high altitudes. The wood has been utilized in building construction, while the resin collected is classified as one of the most valuable Non Timber Forest Products (NTFP) worth up to US $ 50 million/year and applied in many industries. Planting of citron grass (Cymbopogon sp.) has been ’booming’ over the past seven years in the middle regions of Aceh due to its attractive selling price. As consequences, many Pinus trees have been logged and demanded as fire woods in the distillation process of citron grass. Thus, the citron grass’ plantation has been directly threatening the existence of Pinus forests. Therefore, in-situ conservation effort should be started from now on. Here, we would like to report the morphological variation (N= 15 traits) of natural Pinus (e.g., plant height, diameter of main stem, number of crowns per branch, etc.). The morphological data collected were then analysed with the Principal Component Analyses (PCA), in order to cluster the major agronomical parameters among them. This morphological data would be complemented and benefitted the efforts in the genetic study of Aceh Pinus.

DETERMINACIÓN DE LA CONCENTRACIÓN EFECTIVA 50, 90 Y 99 DE IVERMECTINA SOBRE UNA CEPA DE GARRAPATA COMÚN DEL BOVINO (Rhipicephalus (Boophilus) microplus) PROVENIENTE DE LA PROVINCIA DE SANTA FE-ARGENTINA

The aim of this study was to obtain the sensitivity data of the native strains of R. microplus to ivermectin (IVM) in the province of Santa Fe-Argentina, in order to detect the emergence of resistant strains. Native (field isolation) and susceptible (reference) larvae of R. microplus were treated with increasing concentrations of IVM in order to determine its efficacy at different doses. The statistical treatment of the concentration-efficacy curves was performed by the PROBIT model, who is a standardized method proposed by FAO. The results obtained indicated that the larvae isolated were susceptible, being the values of LD50 (9.66 ppm) and LD90 (17.41 ppm), which were lower than those of the reference strain (LD50:13.58 ppm, LD90:19.31 ppm), and the RR (resistance ratio) were less than one (RR50: 0.711; RR90: 0.901). The DL99 (51.48 ppm) was only obtained in the reference strain to determine the discriminant dose (DL99x2), which is a parameter that could be used to easily discriminate susceptible and resistant larvae populations.

Virulence of Beauveria bassiana Strains Isolated from Cadavers of Colorado Potato Beetle, Leptinotarsa decemlineata

The Colorado potato beetle, Leptinotarsa decemlineata (Say), is a serious, widely distributed pest of potato and other crops. This pest is able to defoliate the host plant and cause severe yield loss. Moreover, the pest quickly becomes resistant to many chemical pesticides. Therefore, the development of novel biopesticides targeting this pest is urgently needed. The purpose of this study was to obtain new strains of the entomopathogenic fungus Beauveria bassiana and assess their efficacy against L. decemlineata adults under laboratory conditions. Twelve strains were isolated from cadavers of Colorado potato beetles collected in potato fields in the Czech Republic. Test beetles were treated by suspensions of conidia at the concentration of 1 × 107 spores per milliliter and their survival was recorded daily for three weeks. The results of the bioassays revealed that all new native strains were pathogenic to L. decemlineata adults and caused mortality up to 100% at the end of the trial period with an LT50 of about 7 days. These strains were more virulent than a reference strain GHA and some of them can be recommended for the development of a new mycoinsecticide against L. decemlineata. Our findings also highlight the importance of searching for perspective strains of entomopathogenic fungi among naturally infected hosts.

Soil Microcosms for Bioaugmentation With Fungal Isolates to Remediate Petroleum Hydrocarbon-contaminated Soil

The aim of this work was to isolate indigenous PAH degrading-fungi from petroleum contaminated soil and exogenous ligninolytic strains from decaying-wood, with the ability to secrete diverse enzyme activity. A total of ten ligninolytic fungal isolates and two native strains, has been successfully isolated, screened and identified. The phylogenetic analysis revealed that the indigenous fungi (KBR1 and KB8) belong to the genus Aspergillus niger and tubingensis. While the ligninolytic exogenous PAH-degrading strains namely KBR1-1, KB4, KB2 and LB3 were affiliated to different genera like Syncephalastrum sp, Paecilomyces formosus, Fusarium chlamydosporum, and Coniochaeta sp., respectively. Basis on the taxonomic analysis, enzymatic activities and the hydrocarbons removal rates, single fungal culture employing the strain LB3, KB4, KBR1 and the mixed culture (LB3+KB4) were selected to be used in soil microcosms treatments. The Total petroleum hydrocarbons (TPH), fungal growth rates, BOD5/COD ratios and GC-MS analysis, were determined in all soil microcosmos treatments (SMT) and compared with those of the control (SMU). After 60 days of culture incubation, the highest rate of TPH degradation was recorded in SMT[KB4] by approximately 92±2.35% followed by SMT[KBR1] then SMT[LB3+KB4] with 86.66±1.83% and 85.14±2.21%, respectively.

Trichoderma asperellum (NST-009): A Possible Thai Native Antagonistic Fungus for Managing White Root Disease of Rubber Trees (Hevea brasiliensis)

Background: Rigidoporus microporus causes white root disease, which is one of the most harmful diseases in rubber trees in Thailand. The objectives of this study were to determine the efficacy of T. asperellum NST-009 and its antifungal metabolite in inhibiting R. microporus mycelial development and efficacy of T. asperellum NST-009 in controlling white root disease of rubber trees in an open-field house experiment. Methods: Four native strains of T. asperellum from Nakhon Si Thammarat Province and a commercial strain of Thailand were used in this study. This study was conducted at Agricultural Microbial Production and Service Center, Walailak University, Nakhon Si Thammarat, Thailand, during the period 2017-2020. Result: T. asperellum NST-009 significantly inhibited R. microporus mycelial growth by 77.07% in vitro and its antifungal metabolite from the culture filtrate of T. asperellum NST-009 inhibited mycelial growth by 92.31%. T. asperellum NST-009 reduced the disease severity index by 76.38% in the open-field house experiment compared to the inoculated control. Furthermore, T. asperellum NST-009 was found to survive in rhizosphere soil at 4.50 × 105 CFU/g soil and colonized the roots at 100.00%.

Isolation and identification of probiotic bacteria from Iranian traditional cheese produced by lamb or kid abomasum microorganisms

Different strains of lactic acid bacteria are the most common micro-organisms known as probiotics. Two major kinds of probiotic bacteria are members of the genera Lactobacillus and Bifidobacterium, that are generally associated with the genus Lactobacillus which play a vital role in the body health, as well as the ability to produce antimicrobials and vitamins in the gastrointestinal tract. The purpose of the study was the isolation of probiotics in traditional cheeses that are produced from the natural flora of the newborn mammalian digestive tract. Bacteria from lamb and kid abomasum are added to milk. The curd is slaughtered from the abomasum of a lamb or kid that is suckling and has now no longer began out to devour forage, after which its belly is dried, that is referred to as curd. In this study, sixteen samples of cheeses local to Markazi province of Iran have been prepared. The isolated bacteria have been examined for morphological, biochemical, probiotic properties, and molecular identity. Out of sixteen traditional cheese samples, five kinds of Lactobacillus have been diagnosed through PCR. L. casei strain J026 strain was identified in the traditional cheese samples as the most genera. Using probiotic starters from local sources is a beneficial strategy for producing traditional cheese, which the native strains are more compatible with the humans’ intestinal flora and therefore may also higher play their probiotic’s characteristic.