-
Applied and Environmental Microbiology Sep 2023The application of microfluidic techniques in experimental and environmental studies is a rapidly emerging field. Water-in-oil microdroplets can serve readily as...
The application of microfluidic techniques in experimental and environmental studies is a rapidly emerging field. Water-in-oil microdroplets can serve readily as controllable micro-vessels for studies that require spatial structure. In many applications, it is useful to monitor cell growth without breaking or disrupting the microdroplets. To this end, optical reporters based on color, fluorescence, or luminescence have been developed. However, optical reporters suffer from limitations when used in microdroplets such as inaccurate readings due to strong background interference or limited sensitivity during early growth stages. In addition, optical detection is typically not amenable to filamentous or biofilm-producing organisms that have significant nonlinear changes in opacity and light scattering during growth. To overcome such limitations, we show that volatile methyl halide gases produced by reporter cells expressing a methyl halide transferase (MHT) can serve as an alternative nonoptical detection approach suitable for microdroplets. In this study, an MHT-labeled reporter strain was constructed and characterized. Protocols were established for the encapsulation and incubation of in microdroplets. We observed the complete life cycle for including the vegetative expansion of mycelia, mycelial fragmentation, and late-stage sporulation. Methyl bromide (MeBr) production was detected by gas chromatography-mass spectrometry (GC-MS) from gas reporters incubated in either liquid suspension or microdroplets and used to quantitatively estimate bacterial density. Overall, using MeBr production as a means of quantifying bacterial growth provided a 100- to 1,000-fold increase in sensitivity over optical or fluorescence measurements of a comparable reporter strain expressing fluorescent proteins. IMPORTANCE Quantitative measurement of bacterial growth in microdroplets is desirable but challenging. Current optical reporter systems suffer from limitations when applied to filamentous or biofilm-producing organisms. In this study, we demonstrate that volatile methyl halide gas production can serve as a quantitative nonoptical growth assay for filamentous bacteria encapsulated in microdroplets. We constructed an gas reporter strain and observed a complete life cycle for encapsulated in microdroplets, establishing microdroplets as an alternative growth environment for spp. that can provide spatial structure. We detected MeBr production from both liquid suspension and microdroplets with a 100- to 1,000-fold increase in signal-to-noise ratio compared to optical assays. Importantly, we could reliably detect bacteria with densities down to 10 CFU/mL. The combination of quantitative gas reporting and microdroplet systems provides a valuable approach to studying fastidious organisms that require spatial structure such as those found typically in soils.
Topics: Emulsions; Fluorescence; Gases; Transferases
PubMed: 37699129
DOI: 10.1128/aem.00764-23 -
Insects Aug 2023is a highly polyphagous invasive pest that poses a significant quarantine threat to tropical and subtropical countries. Infested commodities in international trade...
is a highly polyphagous invasive pest that poses a significant quarantine threat to tropical and subtropical countries. Infested commodities in international trade should undergo phytosanitary treatment, and irradiation is recommended as a viable alternative to replace methyl bromide fumigation. Dose-response tests were conducted on the 2-, 4-, and 6-day-old eggs and gravid females of . using the X-ray radiation doses of 15-105 Gy with an interval of 15 Gy. Radiotolerance was compared using ANOVA, fiducial overlapping and lethal dose ratio (LDR) test, resulting in no significant difference among treatments, except for the overall mortality and LDR at LD (a dose causing 90% mortality at 95% confidence level). The estimated dose for LD was 176.5-185.2 Gy, which was validated in the confirmatory tests. No nymphs emerged from a total of 60,386 gravid females exposed to a gamma radiation dose range of 146.8-185.0 Gy in the confirmatory tests. The largest dose in confirmatory tests should be the minimum threshold for phytosanitary treatment, consequently, a minimum dose of 185 Gy is recommended for the phytosanitary irradiation treatment of papaya mealybug-infested commodities, ensuring a treatment efficacy of ≥99.9950% at 95% confidence level.
PubMed: 37623392
DOI: 10.3390/insects14080682 -
Insects Jun 2023The ham mite and the red-legged ham beetle are harmful pests to several high-valued stored products. The regulatory phase-out of the fumigant methyl bromide...
The ham mite and the red-legged ham beetle are harmful pests to several high-valued stored products. The regulatory phase-out of the fumigant methyl bromide necessitates the search for alternative fumigants. Propylene oxide (PPO) and ethyl formate (EF) were therefore evaluated in the laboratory for controlling these pests of dry-cured hams. Concentration-mortality studies at 25 °C of PPO and EF found that the mobile stages of the mites were very susceptible to low concentrations of 10 mg/L and less of each gas, while mite eggs were very tolerant and required 20 mg/L for PPO and 80 mg/L of EF for 100% mortality. Mixed life stage cultures of mites and beetles were treated for 24 h with either PPO or EF at 1× and 2× the estimated 99% lethal doses and confirmed effectiveness for controlling simulated pest populations. The sorptive properties of each gas in chambers with ham pieces, dog food kibbles, or fish meal were minimal for a reduction in mite toxicity when compared to treatments in empty chambers. There was no evidence that any desorbed gas occurred at a level toxic to mite eggs in any of the fumigated commodities. These fumigation studies with ham pests support further work with PPO and EF on any changes in the sensory quality of dry-cured hams for human taste and for commercial-scale fumigations toward regulatory approval.
PubMed: 37367327
DOI: 10.3390/insects14060511 -
Insects May 2023(Hemiptera: Aleyrodidae), commonly known as greenhouse whitefly, is one of the main insect pests of Oriental melon ( var L.) in South Korea. is of concern as a...
(Hemiptera: Aleyrodidae), commonly known as greenhouse whitefly, is one of the main insect pests of Oriental melon ( var L.) in South Korea. is of concern as a quarantine pest for the exportation of in Southeast Asian countries. Due to future restrictions on the use of methyl bromide (MB) during quarantine, ethyl formate (EF) represents a potential alternative. In this study, we evaluated EF for its efficacy (probit-9 values) in enabling the export of Oriental melons. The probit-9 value of EF for controlling was 3.02 g·h/m after 2 h of fumigation. We also assessed the phytotoxicity of EF on melons when using modified atmosphere packaging (MAP) under low-temperature conditions, which is required for export and trade, to extend shelf-life. In scaled-up trials, we found 8 g/m EF for 2 h at 5 °C to be suitable as a new phytosanitary treatment against greenhouse whitefly for exported Oriental melons when using MAP. No phytotoxic damage was found 28 d after fumigation at 5 °C in terms of five quality parameters (firmness, sugar content, mass loss, color change, and external damage).
PubMed: 37233070
DOI: 10.3390/insects14050442 -
Phytopathology Apr 2023Plant pathology plays a critical role in safeguarding plant health, food security, and food safety through science-based solutions to protect plants against recurring...
Plant pathology plays a critical role in safeguarding plant health, food security, and food safety through science-based solutions to protect plants against recurring and emerging diseases. In addition, plant pathology contributed significantly to basic discoveries that have had broad impacts on the life sciences beyond plant pathology. In December 2021, The American Phytopathological Society (APS) conducted a survey among its members and among the readership of its journals to identify and rank key discoveries in plant pathology that have had broad impacts on science and/or practical disease management during the past half century. Based on the responses received, key discoveries that have broadly impacted the life sciences during that period include the Ti plasmid and its mechanism in T-DNA transfer, bacterial ice nucleation, cloning of resistance genes, discovery of viroids, effectors and their mechanisms, pattern-triggered immunity and effector-triggered immunity, RNA interference and gene silencing, structure and function of genes, transcription activator-like effectors, and type-III secretion system and /. Major advances that significantly impacted practical disease management include the deployment and management of host resistance genes; the application of disease models and forecasting systems; the introduction of modern systemic fungicides and host resistance inducers, along with a better understanding of fungicide resistance mechanisms and management; and the utilization of biological controls and suppressive soils, including the implementation of methyl-bromide alternatives. In this special issue, experts from the pertinent fields review the discovery process, recent progress, and impacts of some of the highest ranked discoveries in each category while also pointing out future directions for new discoveries in fundamental and applied plant pathology.
Topics: Plant Pathology; Plant Diseases; Plants; Bacteria; Disease Management; Plant Immunity; Host-Pathogen Interactions
PubMed: 37116465
DOI: 10.1094/PHYTO-02-23-0070-KD -
Insects Mar 2023Sweet persimmons are a valuable export commodity. However, the presence of live insects such as limits their access to many export markets. Methyl bromide,...
Sweet persimmons are a valuable export commodity. However, the presence of live insects such as limits their access to many export markets. Methyl bromide, traditionally used for pest control, is damaging to human health and the environment. Ethyl formate (EF) is a viable alternative; however, its effectiveness against on sweet persimmon fruit is unknown. We evaluated the effectiveness of EF fumigation in controlling present under the calyx of persimmon fruit. The hatching rate of eggs and the survival rates of nymphs and adults of at low temperatures, its LCt and LCt after EF exposure, and phytotoxic damage caused by EF were evaluated in laboratory-scale and commercial-scale tests. The dose-response tests showed that the EF LCt at 5 °C was 9.69, 42.13, and 126.13 g h m for adults, nymphs, and eggs, respectively. Commercial-scale tests demonstrated EF efficacy against all stages without causing phytotoxic effects on persimmons, though the eggs of were not completely controlled in linear low-density polyethylene (LLDPE)-packaged fruit. This study demonstrated that EF is a potential fumigant for quarantine pretreatment, especially before persimmon fruit is packed with LLDPE film, to control infesting sweet persimmon fruit.
PubMed: 37103156
DOI: 10.3390/insects14040341 -
ACS Omega Feb 2023Apple replant disease (ARD) is common in apple production, which seriously affects the growth and development of apples. In this study, hydrogen peroxide with a...
Apple replant disease (ARD) is common in apple production, which seriously affects the growth and development of apples. In this study, hydrogen peroxide with a bactericidal effect was used to treat the replanted soil, and the effects of different concentrations of hydrogen peroxide on replanted seedlings and soil microbiology were investigated in order to seek a green, clean way to control ARD. Five treatments were set up in this study: replanted soil (CK1), replanted soil with methyl bromide fumigation (CK2), replanted soil + 1.5% hydrogen peroxide (H1), replanted soil + 3.0% hydrogen peroxide (H2), and replanted soil + 4.5% hydrogen peroxide (H3). The results showed that hydrogen peroxide treatment improved replanted seedling growth and also inactivated a certain number of , while the , , and also became more abundant in relative terms. The best results were obtained with replanted soil + 4.5% hydrogen peroxide (H3). Consequently, hydrogen peroxide applied to the soil can effectively prevent and control ARD.
PubMed: 36844530
DOI: 10.1021/acsomega.2c06665 -
Plant Disease Feb 2023Globisporangium sylvaticum (syn. Pythium sylvaticum), is an oomycete that causes root rot and damping off of field crops, ornamentals, and vegetables. Several species in...
Globisporangium sylvaticum (syn. Pythium sylvaticum), is an oomycete that causes root rot and damping off of field crops, ornamentals, and vegetables. Several species in Pythiaceae are associated with black root rot of strawberry [(Fragaria × ananassa) Duchesne] (Millner 2006). Mature, stunted 'Chandler' strawberry plants, with reduced shoot vigor and black necrotic roots, were collected from Rhea County (June 2018) and Cumberland County, TN (May 2019). Aboveground symptoms occurred in low incidence (<5% of plants) in the fields. Plant roots were rinsed with tap water, cut into 1 to 3 cm pieces, and surface-disinfested (70% ethanol, 1 min) followed by a sterile water rinse. Root segments were crushed, placed on 20% V8 juice agar, and incubated in the dark at 21°C for 3 days. White fluffy mycelia grew from a majority of roots and coenocytic hyphae with globose hyphal swellings, delimited from hyphae by septa, were observed with microscopy. Hyphae were initially branched, curled, hyaline, and aseptate; however, septations were observed in older cultures. Globose structures (terminal and intercalary) were identified as sporangia [11 to 32 (avg. 22.1) µm diameter] when zoospores were observed (Parikh et al. 2022). Oospores [9 to 21 (avg. 16) μm diameter] were globose, smooth, aplerotic, and thick-walled. Oogonia, with or without one or more inflated antheridia, were observed when isolates were paired in culture, characteristics consistent with descriptions of Campbell and Hendrix (1967), Pratt and Green (1971), van der Plaats-Niterink (1981), and Uzuhashi et al. (2010). Genomic DNA was extracted (Extract-N-Amp™; Sigma-Aldrich, MO) for PCR amplification of internal transcribed spacer (ITS) regions of rDNA with primers ITS1/ITS4 (White et al. 1990); ITS and large subunit rRNA regions with primers UN-up18S42/UN-lo28S22 (Robideau et al. 2011); and cytochrome c oxidase subunit I (COI) mitochondrial DNA with primers OomCoxI-Levup/OomCoxI-Levlo (Robideau et al. 2011). Primers ITS1/ITS4 were used to amplify isolate TN (GenBank Accession MW386310, which had 100% homology with reference isolate MK326528). Primers UN-up18S42/UN-lo28S22 amplified isolates SAP18 and OO1 (Accessions MZ881935 and MZ881936, which had 99.8% homology with HQ665236), and COI primers amplified isolate SAP18 (Accession OK020192, which had 100% homology with GU071816 and KT692835). To satisfy Koch's postulates, inoculum of G. sylvaticum grown on autoclaved wheat seeds was added (5% w/v) to planting mix (1 peat:1 sand, v/v). Young, rooted strawberry plants were planted in 1.2-L pots with infested (n = 6) and control (no pathogen, n = 6) mixes, which was saturated with deionized water. Pots were covered with clear plastic for 48 h to maintain high humidity. Plants were grown in a greenhouse (24°C avg.) for 8 weeks. The disease assay was repeated. All plants in infested mix died, with black, necrotic roots. Plants in the control mix were healthy and well-established. The pathogen was reisolated from roots of all inoculated plants and confirmed to be G. sylvaticum based on morphology and molecular analyses. Root disease of strawberry caused by G. sylvaticum has been reported in the USA (Campbell and Hendrix 1967; Nemec and Sanders 1970; Pratt and Green 1971). This is the first report of G. sylvaticum causing root rot of strawberry in Tennessee. With the loss of methyl bromide, sustainable disease control strategies are needed to provide effective management options for strawberry black root rot.
PubMed: 36825322
DOI: 10.1094/PDIS-01-23-0007-PDN -
Scientific Reports Feb 2023Fusarium wilt, the most serious soil-borne pathogen, is a serious problem for tomato production worldwide. The presented study evaluated the antifungal activity against...
Fusarium wilt, the most serious soil-borne pathogen, is a serious problem for tomato production worldwide. The presented study evaluated the antifungal activity against Fusarium oxysporum f. sp. lycopersici in vitro and in vivo for nine fumigants. In addition, the research examined the possibility of enhancing the growth of tomato plants in order to increase resistance against this disease by using four chemical inducers. The results indicated that at 20 mg/L, the radial growth of the pathogen was inhibited 100% by formaldehyde and > 80% by phosphine. Among the essential oils investigated, neem oil was the most effective, however, it only achieved 40.54% at 500 mg/L. The values of EC for all fumigants, except dimethyl disulfide (DMDS) and carbon disulfide (CS2), were lower than those for thiophanate-methyl. Phosphine was the highest efficient. The elicitors can be arranged based on their effectiveness, gibberellic acid (GA3) > sorbic acid > cytokinin (6-benzylaminopurine) > indole-3-butyric acid. The change in root length, fresh weight, and dry weight was greater with soil drench than with foliar application. The fumigant generators formaldehyde, phosphine and 1,4-dichlorobenzene and bio-fumigants citrus and neem oils as well as elicitors gibberellic and sorbic acid could be one of the promising alternatives to methyl bromide against Fusarium oxysporum as an important component of integrated management of Fusarium wilt.
Topics: Fusarium; Solanum lycopersicum; Sorbic Acid; Pesticides; Soil; Plant Diseases
PubMed: 36774421
DOI: 10.1038/s41598-023-29033-w -
Sangyo Eiseigaku Zasshi = Journal of... May 2023The studies on occupational peripheral nerve disorders (OPD) in the workplace and future issues of OPD were overviewed based on the author's experiences.
OBJECTIVES
The studies on occupational peripheral nerve disorders (OPD) in the workplace and future issues of OPD were overviewed based on the author's experiences.
METHODS
The author shows four case studies because of four chemicals; cross-sectional studies using nerve conduction velocity (NCV) on the effect on the peripheral nerve among workers exposed to carbon disulphide (CS) in rayon manufacturing factories and studies on OPD among patients with vibration syndrome (VS) and non-specific cervicobrachial disorders (CBD).
RESULTS
Four cases of OPD due to chronic poisoning because of inorganic lead, thallium, n-hexane, and methyl bromide, respectively, were presented with clinical signs and symptoms of OPD and the cause-and-effect relationships were described. Cross-sectional epidemiological studies clarified that workers exposed to approximately 5 ppm of CS showed NCV reduction in the lower extremities and those exposed to > 5 ppm showed NCV reduction in the upper extremities. Multifocal damages due to VS was presented through high rate of patients with sensory nerve conduction velocity (SCV) reduction in multiple nerves. SCV reduction of the radial nerve in the index finger among patients with CBD was presented.
CONCLUSIONS
The various OPD phases were shown by case, patient, and epidemiological study. However, the recent decrease of work-related illness due to chemicals may result to OPD reduction. The issue of OPD including other factors and differential diagnosis, will remain eventually owing to introducing chemicals of which toxicity are unclear.
Topics: Humans; Neural Conduction; Cross-Sectional Studies; Peripheral Nervous System Diseases; Peripheral Nerves; Occupational Diseases
PubMed: 36754411
DOI: 10.1539/sangyoeisei.2022-036-A