Evaluation of ultra-low volume treatments on Culex quinquefasciatus (Diptera: Culicidae) abundance and relative age by parity dissection.

St. Tammany Parish Mosquito Abatement District relies on ultra-low volume (ULV) insecticides to manage adult Culex quinquefasciatus (Say 1823), the primary vector of West Nile virus in Louisiana, but abundance rarely declines post-treatment. Three field ULV applications each of adulticides containing deltamethrin, naled, or resmethrin were conducted from May through October 2022 to measure efficacy against free-flying Cx. quinquefasciatus. Female mosquitoes trapped before and after treatment were identified, counted, and age-graded by parity dissection. No insecticide applications reduced Cx. quinquefasciatus abundance, but trials with deltamethrin and naled significantly lowered average mosquito age in the area post-treatment. No trials with resmethrin reduced mosquito age, which corroborates laboratory susceptibility data indicating local resistance toward the pyrethroid. These trials establish parity dissections and relative age gradation as a means of measuring treatment success against Cx. quinquefasciatus.

Field Assessment of Naled and Its Primary Degradation Product (Dichlorvos) in Aquatic Ecosystems Following Aerial Ultra-low Volume Application for Mosquito Control.

Naled, an organophosphate insecticide, is applied aerially at ultra-low volumes over aquatic ecosystems near Sacramento, California, USA, during summer months for mosquito control. Two ecosystem types (rice fields and a flowing canal) were sampled in 2020 and 2021. Naled and its primary degradation product (dichlorvos) were measured in water, biofilm, grazer macroinvertebrates, and omnivore/predator macroinvertebrates (predominantly crayfish). Maximum naled and dichlorvos concentrations detected in water samples one day after naled application were 287.3 and 5647.5 ng/L, respectively, which were above the U.S. Environmental Protection Agency's aquatic life benchmarks for invertebrates. Neither compound was detected in water more than one day after the application. Dichlorvos, but not naled, was detected in composite crayfish samples up to 10 days after the last aerial application. Detections in water from the canal showed that the compounds were transported downstream of the target application area. Factors such as vector control flight paths, dilution, and transport through air and water likely affected concentrations of naled and dichlorvos in water and organisms from these aquatic ecosystems.

Photolytic degradation of novel polymeric and monomeric brominated flame retardants: Investigation of endocrine disruption, physiological and ecotoxicological effects.

Ecotoxicological effects of photolytic degradation mixtures of the two brominated flame retardants PolymericFR and Tetrabromobisphenol A-bis (2,3-dibrom-2-methyl-propyl) Ether (TBBPA-BDBMPE) have been studied in vitro and in vivo. Both substances were experimentally degraded separately by exposure to artificial UV-light and the resulting degradation mixtures from different time points during the UV-exposure were applied in ecotoxicological tests. The in vitro investigation showed no effects of the degraded flame retardants on the estrogenic and androgenic receptors via the CALUX (chemically activated luciferase gene expression) assay. Short-term exposures (up to 96 h) of Lumbriculus variegatus lead to temporary physiological reactions of the annelid. The exposure to degraded PolymericFR lead to an increased activity of Catalase, while the degradation mixture of TBBPA-BDBMPE caused increases of Glutathione-S-transferase and Acetylcholine esterase activities. Following a chronic exposure (28 d) of L. variegatus, no effects on the growth, reproduction, fragmentation and energy storage of the annelid were detected. The results indicate that the experimental degradation of the two flame retardants causes changes in their ecotoxicological potential. This might lead to acute physiological effects on aquatic annelids, which, however, do not affect the animals chronically according to our results.

Interactions of organophosphorus pesticides with ATP-binding cassette (ABC) drug transporters.

Although pharmaceutical companies have to study drug-transporter interaction, environmental contaminant interactions with these transporters are not well characterised. In this study, we demonstrated using in vitro transfected cell line that some organophosphorus pesticides are able to interact with drug efflux transporters like P-glycoprotein, BCRP and MRPs.According to our results, dibrom was found to inhibit only Hoechst binding site of P-gp with an IC50 closed to 77 µM, phosmet inhibited BCRP efflux with an IC50 of 42 µM and only profenofos was able to inhibit BCRP, MRPs and P-gp at two binding sites. As profenofos appeared to be a potent ABC transporter inhibitor, we studied its potential substrate property towards P-gp.Using a docking approach, we developed an in silico tool to study pesticide properties to be a probe or inhibitor of P-gp transporter. From both in silico and in vitro results, profenofos was not considered as a P-gp substrate.Combining both in vitro and docking methods appears to be an attractive approach to select pesticides that would not pass into the blood systemic circulation.

Insecticide Susceptibility Status of Lab and Field Populations of Aedes albopictus from Gainesville, Florida, to Organophosphates and Pyrethroids.

Previous unpublished results from 2 open-field studies in South Florida evaluated the effectiveness of 4 ultra-low volume malathion formulations, applied by ground application against a USDA (Gainesville) laboratory-colonized strain of Aedes albopictus. Field results of the bioassays indicated that the species was highly resistant to malathion compared with other insecticide-susceptible mosquito species concurrently tested. Unfortunately, when the Ae. albopictus colony was initially started in 2003 from a field collection in Gainesville, FL, susceptibility to insecticides was not determined. Since that time, this species has been in constant production without exposure to pesticides by the Center for Medical, Agricultural, and Veterinary Entomology, Gainesville, FL. In order to determine its susceptibility level to malathion and compare it with 3 current field-collected Ae. albopictus populations, 3 organophosphates (malathion, chlorpyrifos, naled) and 2 pyrethroids (permethrin, deltamethrin) were tested in the laboratory, using the Centers for Disease Control and Prevention bottle bioassays. Results of the study confirmed that the USDA-Gainesville Ae. albopictus was still highly resistant to malathion even after being maintained in the laboratory for 16 years without exposure to pesticides. The 3 field-collected populations were also resistant to malathion but slightly less so than the USDA-Gainesville population. Rotation of pyrethroid products 16 years ago did not result in any significant reduction of malathion resistance in the field populations. Neither the USDA-Gainesville nor the field-collected Ae. albopictus populations showed cross-resistance to chlorpyrifos and naled. Lower mortalities were observed in the 10-min diagnostic time from the 3 field-collected populations against permethrin but rose significantly at 15-min exposure. Resistance was not detected for deltamethrin in any of the 4 Gainesville Ae. albopictus populations.

Persistence of aerially-sprayed naled in coastal sediments.

Aerial sprays of the organophosphate pesticide, naled, were intensified over beach areas during the summer of 2016 to control the locally-acquired Zika outbreak in the continental U.S. Concerns were raised in beach frequented areas about contaminated sediments. The aim of this study was to evaluate the persistence and levels of naled and its byproduct, dichlorvos, in sediments obtained from the affected areas. Laboratory experiments were designed to simulate the effect of various natural conditions on the decomposition of naled in three sediment types (beach sand, marl, and calcinated beach sand). The three sediment samples were also exposed to field aerial sprays. After 30 min of exposure, more dichlorvos was detected in the sediments than naled, with 33 to 43% of the molar concentration initially applied as either naled or dichlorvos. Under dark conditions, trace levels of naled were observed after 24 h on sediments. Higher temperature accelerated the natural decomposition of both naled and dichlorvos in sediments. The half-life of naled ranged from 3 to 5 h at 22.5 °C and ranged from 1 to 3 h at 30 °C. Expedited decomposition of naled was observed under sunlight conditions with a half-life of naled of 20 min. In the field, only dichlorvos was detected in the sediment samples at concentrations between 0.0011 and 0.0028 µmol/g 1 h after aerial sprays. This data can be used towards a risk assessment that evaluates exposures to naled and dichlorvos through beach sands impacted by aerial spray activities.

Mortality and Cholinesterase Inhibition in Butterflies Following Aerial Naled Applications for Mosquito Control on the National Key Deer Refuge.

Natural resource managers are concerned about the impacts of aerial ultra-low volume spray (ULV) of insecticides for mosquito control (i.e., mosquito adulticides) and seek science-driven management recommendations that reduce risk but allow vector control for nearby human populations. Managers at the National Key Deer Refuge (Florida Keys, FL) are concerned for ULV effects upon conservation efforts for imperiled butterflies (Florida leafwing [Anaea troglodyta floridalis] and Bartram's hairstreak [Strymon acis bartrami] butterflies). No-spray zones were designated for protection of those butterflies, but their effectiveness for mitigation is unclear. To address this uncertainty, cholinesterase activity (ChE) and mortality were monitored for caged butterflies gulf fritillary [Agraulis vanilla] and great southern white [Ascia monuste]) deployed on the Refuge during three aerial ULV applications of the insecticide naled. Residue samplers also were deployed to estimate butterfly exposure. Spray efficacy against mosquitoes was assessed by deploying caged mosquitoes at the same locations as the butterflies. Average naled residue levels on filter paper samplers in the target area (1882-2898 µg/m2) was significantly greater than in the no-spray zone (9-1562 µg/m2). Differences between the no-spray zone and target area for butterfly mortality and ChE were inconsistent. Average mortality was significantly lower, and average ChE was significantly higher in the no-spray zone for larvae of one species but not for larvae of the other species. Mosquito mortality did not differ significantly between the two areas. Data from the present study reflect the inconsistent effectiveness of no-spray zones on the Refuge using standard methods employed at the time by the vector control agency in the Florida Keys and possibly by other vector control agencies in similar coastal environments. Furthermore, these findings helped to guide the design and to improve the conservation value of future no-spray zone delineations while allowing for treatment in areas where mosquito control is necessary for vector-borne disease reduction.

Microbial communities from subglacial water of naled ice bodies in the forefield of Werenskioldbreen, Svalbard.

We assessed the structure of microbial communities in the subglacial drainage system of the Werenskioldbreen glacier, Svalbard, which consists of three independent channels. Dome-shaped naled ice bodies that had been forming and releasing subglacial water in the glacial forefield during accumulations season were used to study glacial microbiome. We tested the hypothesis that the properties of the water transported by these channels are site-dependent and influence bacterial diversity. We therefore established the phylogenetic structure of the subglacial microbial communities using next generation sequencing (NGS) of the 16S rRNA gene and performed bioinformatics analyses. A total of 1409 OTUs (operational taxonomic units) belonged to 40 phyla; mostly Proteobacteria, Gracilibacteria, Bacteroidetes, Actinobacteria and Parcubacteria were identified. Sites located on the edge of Werenskioldbreen forefield (Angell, Kvisla) were mainly dominated by Betaproteobacteria. In the central site (Dusan) domination of Epsilonproteobacteria class was observed. Gracilibacteria (GN02) and Gammaproteobacteria represented the dominant taxa only in the sample Kvisla 2. Principal Coordinate Analysis (PCoA) of beta diversity revealed that phylogenetic profiles grouped in three different clusters according to the sampling site. Moreover, higher similarity of bacterial communities from Angell and Kvisla compared to Dusan was confirmed by cluster analysis and Venn diagrams. The highest alpha index values was measured in Dusan. Richness and phylogenetic diversity indices were significantly (p < .05) and positively correlated with pH values of subglacial water and negatively with concentration of Cl-, Br-, and NO3- anions. These anions negatively impacted the values of richness indices but positively correlated with abundance of some microbial phyla. Our results indicated that subglacial water from naled ice bodies offer the possibility to study the glacial microbiome. In the studied subglacial water, the microbial community structure was sampling site specific and dependent on the water properties, which in turn were probably influenced by the local bedrock composition.

Meeting Increased Demand for Mosquito Adulticides Containing the Active Ingredient Naled Following Hurricanes and Tropical Storms.

The occurrence of tropical storms and hurricanes is a certainty in the Atlantic Basin each year. Many of these never make landfall. Those that do can range in intensity from a weak tropical depression to a very destructive Category 5 hurricane. These storms often produce large amounts of rainfall and flooding, resulting in increases in the mosquito populations in the affected areas. In order to deal with this problem, aerial applications of insecticides over wide areas can provide relief to the impacted area, assisting in the recovery efforts. Meeting the demand for these sudden and large increases in the volume of the insecticide most commonly used in aerial applications requires great coordination, communication, and commitment. We describe the diverse entities involved in the manufacture, distribution, and use of the product and how this increase in need is recognized, managed, and satisfied in a compressed period of time.

Texas Mosquito Control Response Following Hurricane Harvey.

Hurricane Harvey has been recorded as the wettest cyclone in United States history, resulting in devastating and catastrophic flooding for the Texas Gulf Coast. The nature of the path of the hurricane, with multiple landfalls along the Texas Gulf Coast, resulted in the largest aerial mosquito control effort for one single storm. Two mosquito control contractors and the Air Force Aerial Spray Unit of the US Air Force Reserve were used to aerial treat 6,765,971 acres (3,075,441 ha) in 29 of the 60 disaster-declared counties in Texas. During the response, 101,253 liters of Dibrom® (active ingredient [AI]: naled) and 48,735 liters of Duet™ (AI: 1% prallethrin and 5% sumithrin) were used. In 23/29 counties requesting aerial spraying, mosquito control contractors were used to conduct pre- and postaerial application mosquito surveillance. The remaining 6 counties conducted their own surveillance during the response. A total of 105,153 mosquitoes in 7 genera and 35 species were collected during this response with the major floodwater nuisance mosquito being Psorophora columbiae. The most abundant vector mosquito collected was Culex nigripalpus. Duet at the 0.8% and 1% application rates resulted in 49% and 69% control of Ps. columbiae, respectively. Dibrom application resulted in 95% and 93% control of Ps. Columbia and Cx. nigripalpus populations, respectively.

Nighttime Aerial Sprays for Control of Crepuscular Biting Midges in South Carolina.

Nighttime aerial spray applications with naled were conducted to evaluate their efficacy in controlling crepuscular biting midges (Culicoides spp.) in South Carolina, using a US Air Force C-130. Local populations of Culicoides spp. were monitored before and after the sprays with Mosquito Magnet traps to assess the efficacy of postsunset applications. Biting midge populations were consistently decreased by the aerial spray applications in this study. This indicates that nighttime sprays can be used to control these pests, even when their peak flight activity is focused around sunset.

Bacteria-mediated modification of insecticide toxicity in the yellow fever mosquito, Aedes aegypti.

The incidence of mosquito-borne disease poses a significant threat to human and animal health throughout the world, with effective chemical control interventions limited by widespread insecticide resistance. Recent evidence suggests that gut bacteria of mosquitoes, known to be essential in nutritional homeostasis and pathogen defense, may also play a significant role in facilitating insecticide resistance. This study investigated the extent to which bacteria contribute to the general esterase and cytochrome P450 monooxygenase (P450)-mediated detoxification of the insecticides propoxur and naled, as well as the insecticidal activity of these chemistries to the yellow fever mosquito, Aedes aegypti. Experiments conducted using insecticide synergists that reduce general esterase and P450 activity demonstrate a role for both groups of enzymes in the metabolic detoxification of propoxur and naled. Furthermore, reduction of bacteria in mosquito larvae using broad-spectrum antibiotics was found to decrease the metabolic detoxification of propoxur and naled, suggesting that the bacteria themselves may be contributing to the in vivo metabolic detoxification of these insecticides. This was supported by in vitro assays using culturable gut bacteria isolated from mosquito larvae which demonstrated that the bacteria were capable of reducing insecticide toxicity. More work is needed, however, to fully elucidate the contribution of bacteria in Ae. aegypti larvae to the metabolic detoxification of insecticides.

Rapid Screening of Aedes aegypti Mosquitoes for Susceptibility to Insecticides as Part of Zika Emergency Response, Puerto Rico.

In response to the 2016 Zika outbreak, Aedes aegypti mosquitoes from 38 locations across Puerto Rico were screened using Centers for Disease Control and Prevention bottle bioassays for sensitivity to insecticides used for mosquito control. All populations were resistant to pyrethroids. Naled, an organophosphate, was the most effective insecticide, killing all mosquitoes tested.

Detection of Male Oriental Fruit Fly (Diptera: Tephritidae) Susceptibility to Naled- and Fipronil-Intoxicated Methyl Eugenol.

Naled-intoxicated methyl eugenol (ME) is commonly used to control oriental fruit flies, Bactrocera dorsalis (Hendel) (Diptera: Tephritidae), in Taiwan. However, non-responsiveness to ME and pesticide resistance in oriental fruit flies may reduce control efficacy. In this study, mark-recapture experiments were used to analyze the effects of naled-intoxicated ME on field and naled-resistant fly strains. ME non-responsiveness was tested in field, naled-resistant, and susceptible strains and pyrosequencing techniques were used to detect frequencies of point mutations on organophosphate resistant alleles in field strains. Finally, the effects of fipronil-intoxicated ME were analyzed to determine whether control efficiency could be enhanced through the use of alternate pesticides. Control efficiency of naled-intoxicated ME was found to be significantly lower in the field and resistant strains compared to the susceptible strain. ME non-responsiveness was found to be 1.7-1.9% in our lab-reared strains (both naled-resistant and susceptible) and 3.4-4.3% in field strains. Results of our pyrosequencing study found frequency of resistant alleles in captured male field flies to be significantly lower than that of the original population, indicating that it is highly probable that resistant flies may escape from traps. Finally, capture rates of naled-resistant flies increased when naled was replaced with fipronil in attractants, showing that use of pesticides with different modes of action could possibly increase control efficiency of intoxicated ME attractants.

Elemental imaging (LA-ICP-MS) of zebrafish embryos to study the toxicokinetics of the acetylcholinesterase inhibitor naled.

The zebrafish embryo is an important model in ecotoxicology but the spatial distribution of chemicals and the relation to observed effects is not well understood. Quantitative imaging can help to gain insights into the distribution of chemicals in the zebrafish embryo. Laser ablation inductively coupled plasma-mass spectrometry (LA-ICP-MS) is used to quantify the uptake and the uptake kinetics of the bromine (Br) containing organophosphate naled (Dibrom®, dimethyl-1,2-dibromo-2,2-dichloroethylphosphate) and its distribution in zebrafish embryos using Br as the marker element. During exposure, the Br amounts increase in the embryos parallel to the irreversible inhibition of the acetylcholinesterase (AChE). The final amount of Br in the embryo (545 pmol/embryo) corresponds to a 280-fold enrichment of naled from the exposure solution. However, LC-MS/MS analyses showed that the internal concentration of naled remained below the LOD (7.8 fmol/embryo); also the concentration of its known transformation product dichlorvos remained low (0.85 to 2.8 pmol/embryo). These findings indicate the high reactivity and high transformation rate of naled to other products than dichlorvos. 12C normalized intensity distributions of Br in the zebrafish embryo showed an enrichment of Br in its head region. Kernel density estimates of the LA-ICP-MS data were calculated and outline the high reproducibility between replicated and the shift in the Br distribution during exposure. The Br enrichment indicates a preferential debromination or direct covalent reaction of naled with AChE in this region. Graphical abstract ᅟ.

Poor qualitative analysis of Naled necessarily leads to incorrect quantitative analysis.

This correspondence is in regards to a published article, titled "Prenatal Naled and Chlorpyrifos Exposure is Associated with Deficits in Infant Motor Function in a Cohort of Chinese Infants" Silver et al., Environ Int., 2017 Sep; 106:248-256. Upon careful review of this work we identified some significant issues in the mass spectral analysis of naled, specifically its identification and quantification. In this communication we address these issues and provide analytical data and rationale to support our criticism of the reported work. We collected mass spectral data for naled (analytical standard grade) under a variety of mass spectrometric conditions in an attempt to obtain a fragmentation pattern similar to what was reported in Silver et al., 2017. We however, could not reproduce a similar fragmentation pattern under any of the tested experimental conditions. Our results however were in excellent agreement with the National Institute of Standards and Technology (NIST) database, which is a very well established and widely accepted resource for such compounds. This leads us to conclude that naled was in all probability misidentified in the reported (Silver et al., 2017) study which consequently raises serious questions regarding the quantification of naled in the blood samples thus placing the whole statistical correlation of naled as a contributor to impairment of motor function in infants in question.

Effects of Mosquito Control Adulticides on Sterile Cochliomyia hominivorax (Diptera: Calliphoridae).

Effects of mosquito control adulticides on sterile screwworm flies, Cochliomyia hominivorax (Coquerel) (Diptera: Calliphoridae), were investigated via bottle bioassays, outdoor cage tests, and exposure to treated vegetation. In bottle bioassays, 43 µg of permethrin via dilution of Evoluer, 474.56 µg of malathion via dilution of Fyfanon, and 25 µg of naled via dilution of Dibrom Concentrate were used to challenge screwworm flies. Permethrin was more toxic to screwworm flies than was malathion, which was more toxic than naled. On succeeding days, permethrin was still lethal to the flies, whereas malathion and naled were less toxic. During outdoor cage trials, screwworm mortality declined as distance from the spray truck increased. Sterile screwworm flies were killed by lower concentrations of permethrin needed to kill black salt marsh mosquitoes, Aedes taeniorhynchus (Wiedemann) (Diptera: Culicidae). Flies exposed to treated vegetation taken from the path of the spray cloud died more quickly than did flies exposed to leaves taken 5 ft inside the canopy. Fly mortality increased as volume mean diameter of droplets increased. In spite of the toxicity of Evoluer to screwworm flies, aspects of their biology make it unlikely that mosquito control operations would affect released flies.

Aerial ULV control of Aedes aegypti with naled (Dibrom) inside simulated rural village and urban cryptic habitats.

We conducted aerial fixed wing ultra low volume (ULV) spray trials with naled to investigate penetration of exposed and simulated cryptic habitat within opened buildings, partially sealed buildings, and outdoor locations targeting sentinel adult Aedes aegypti mosquitoes in north central Florida. Mortality was observed in open and closed buildings and outdoors, even in mosquitoes placed in cryptic habitats. Observations on the impact of building type, mosquito exposure method such as placement in cryptic habitat, and spray nozzle size on mosquito mortality are described and analyzed.

Prenatal naled and chlorpyrifos exposure is associated with deficits in infant motor function in a cohort of Chinese infants.

BACKGROUND: Organophosphate insecticides (OPs) are used worldwide, yet despite nearly ubiquitous exposure in the general population, few have been studied outside the laboratory. Fetal brains undergo rapid growth and development, leaving them susceptible to long-term effects of neurotoxic OPs. The objective here was to investigate the extent to which prenatal exposure to OPs affects infant motor development. METHODS: 30 OPs were measured in umbilical cord blood using gas chromatography tandem mass spectrometry in a cohort of Chinese infants. Motor function was assessed at 6-weeks and 9-months using Peabody Developmental Motor Scales 2nd edition (PDMS-2) (n=199). Outcomes included subtest scores: reflexes, stationary, locomotion, grasping, visual-motor integration (V-M), composite scores: gross (GM), fine (FM), total motor (TM), and standardized motor quotients: gross (GMQ), fine (FMQ), total motor (TMQ). RESULTS: Naled, methamidophos, trichlorfon, chlorpyrifos, and phorate were detected in ≥10% of samples. Prenatal naled and chlorpyrifos were associated with decreased 9-month motor function. Scores were 0.55, 0.85, and 0.90 points lower per 1ng/mL increase in log-naled, for V-M (p=0.04), FM (p=0.04), and FMQ (p=0.08), respectively. For chlorpyrifos, scores were 0.50, 1.98, 0.80, 1.91, 3.49, 2.71, 6.29, 2.56, 2.04, and 2.59 points lower for exposed versus unexposed infants, for reflexes (p=0.04), locomotion (p=0.02), grasping (p=0.05), V-M (p<0.001), GM (p=0.007), FM (p=0.002), TM (p<0.001), GMQ (p=0.01), FMQ (p=0.07), and TMQ (p=0.008), respectively. Girls appeared to be more sensitive to the negative effects of OPs on 9-month motor function than boys. CONCLUSIONS: We found deficits in 9-month motor function in infants with prenatal exposure to naled and chlorpyrifos. Naled is being aerially sprayed to combat mosquitoes carrying Zika virus, yet this is the first non-occupational human study of its health effects. Delays in early-motor skill acquisition may be detrimental for downstream development and cognition.