Search

Since the US Food and Drug Administration's approval of monensin in 2004, significant nutritional advances have been made to increase feed efficiency and milk fat production. Recent evidence suggests monensin's adverse effect on milk fat percentage may be absent when diets are formulated to address known diet-induced milk fat depression risk factors. Thus, study objectives were to evaluate effects of monensin level on dry matter intake (DMI), milk production and composition, and efficiency of high-producing cows fed diets formulated to optimize milk fat. Ninety-six lactating Holstein cows (36 primiparous, 60 multiparous; 106 ± 17 d in milk [DIM]) were balanced by parity, DIM, and milk production and were randomly assigned to 1 of 12 pens with 8 cows per pen. All cows received 11 g/t monensin for 5 wk after which pens received 1 of 4 dietary treatments (n = 3) formulated to provide 0 (CON), 11 (R11), 14.5 (R14.5), or 18 (R18) g/t monensin for 9 wk. The basal diet was 54% forage, 27% NDF, 29% starch, and 2.3% rumen unsaturated fatty acid load. Pen was the experimental unit and data were analyzed using the Fit Model Procedure of JMP. Effects of treatment, time, and treatment × time interaction were included as fixed effects and pen as a random effect. Least squares means were determined and linear and quadratic contrasts were tested. Dry matter intake tended to decrease linearly with increasing monensin dose. Milk yield, fat percentage, and protein percentage and yield were unaffected by treatment while fat yield was quadratically increased. Milk de novo and mixed fatty acid (FA) yields (g/d) increased quadratically with monensin whereas preformed FA linearly decreased during the experimental period. Energy-corrected milk (ECM) was quadratically increased by monensin. Milk urea nitrogen concentrations increased linearly with increasing monensin dose. Monensin linearly increased feed efficiency (ECM/DMI, 3.5% fat-corrected milk/DMI, and solids-corrected milk/DMI). Body weight gain did not differ between treatments. Estimated dietary energy tended to increase linearly with increasing monensin level. These data suggest monensin improves component-corrected milk production efficiency, estimated dietary energy, and does not negatively affect milk fat percentage or FA profile., (The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Our objectives were to evaluate the effects of prepartum monensin supplementation and dry-period nutritional strategy on the postpartum productive performance of cows fed monensin during lactation. A total of 102 Holstein cows were enrolled in the experiment (32 primiparous and 70 multiparous). The study was a completely randomized design, with randomization restricted to balance for parity, body condition score, and expected calving date. A 2 × 2 factorial arrangement of prepartum treatments was used; the variables of interest were prepartum feeding strategy [controlled-energy diet throughout the dry period (CE) vs. controlled-energy diet from dry-off to 22 d before expected parturition, followed by a moderate-energy close-up diet from d 21 before expected parturition through parturition (CU)] and prepartum monensin supplementation [0 g/t (control, CON) or 24.2 g/t (MON); Rumensin; Elanco Animal Health, Greenfield, IN]. Lactation diets before and after the dry period contained monensin at 15.4 g/t. During the close-up period, cows fed CU had greater DM and NE L intakes than cows fed CE. Calf BW at birth tended to be greater for cows fed CU than for those fed CE but was not affected by MON supplementation. Diet did not affect calving difficulty score, but cows supplemented with MON had an increased calving difficulty score. We found a tendency for a MON × parity interaction for colostral IgG concentration, such that multiparous MON cows tended to have lower IgG concentration than CON cows, but colostral IgG concentration for primiparous MON and CON cows did not differ. Postpartum milk yield did not differ between diets but tended to be greater for cows supplemented with MON. Milk fat and lactose content were greater for cows fed CU than for those fed CE, and lactose content and yield were increased for cows supplemented with MON. Solids-corrected and fat-corrected milk yields were increased by MON supplementation, but were not affected by diet. Overall means for postpartum DMI did not differ by diet or MON supplementation. The CU diet decreased the concentration of nonesterified fatty acids during the close-up period but increased it postpartum. Neither diet nor monensin affected β-hydroxybutyrate or liver composition. Overall, postpartum productive performance differed little between prepartum dietary strategies, but cows fed MON had greater energy-corrected milk production. In herds fed monensin during lactation, monensin should also be fed during the dry period., (Copyright © 2021 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Nonclinical testing of human pharmaceuticals is conducted to assess the safety of compounds to be studied in human clinical trials and for marketing of new drugs. Although there is no exact number and type of nonclinical studies required for safety assessments, as there is inherent flexibility for each new compound, the traditional approach is outlined in various FDA and ICH guidance documents and involves a combination of in vitro assays and whole animal testing methods. Recent advances in science have led to the emergence of numerous new approach methodologies (NAMs) for nonclinical testing that are currently being used in various aspects of drug development. Traditional nonclinical testing methods can predict clinical outcomes, although improvements in these methods that can increase predictivity of clinical outcomes are encouraged and needed. This paper discusses FDA/CDER's view on the opportunities and challenges of using NAMs in drug development especially for regulatory purposes, and also includes examples where NAMs are currently being used in nonclinical safety assessments and where they may supplement and/or enhance current testing methods. FDA/CDER also encourages communication with stakeholders regarding NAMs and is committed to exploring the use of NAMs to improve regulatory efficiency and potentially expedite drug development., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The hurricane is no stranger to longtime residents of Florida's east coast. In 1979, after about 15 years of local inactivity, Hurricane David made landfall in West Palm Beach. Thirteen years later and 100 miles south, category 5 Hurricane Andrew caused catastrophic damage when it hit the city of Homestead in the Miami-Dade area. In 2004, the counties along the east coast of central Florida were hit by 2 devastating hurricanes, Frances and Jeanne, that made landfall at Sewall's Point just 20 days apart. The very next year, Hurricane Wilma made landfall near Everglades City as a Category 3 storm. After a decade of relief, a glancing blow from Hurricane Matthew struck in 2016, only to be followed by the extremely devastating Hurricane Irma just 1 year later. Each of these hurricanes caused significant property damage and mosquito problems for the Florida residents affected by these storms. In 1997, the Indian River Mosquito Control District (IRMCD) developed a hurricane preparedness plan outlining the appropriate action to be taken depending on the severity of the approaching storm. The IRMCD has also learned to negotiate the intricacies of the Federal Emergency Management Agency's reimbursement program, thus reducing the financial impact to the District. This paper provides an overview of how IRMCD has prepared, reacted, and followed-up with the seemingly constant parade of hurricanes that have threatened and affected the east coast over time., (Copyright © 2020 by The American Mosquito Control Association, Inc.)

The aims of this review were to compare planning for both mosquito control and land use in east-central Florida, USA, and in New South Wales, Queensland, and the Northern Territory, Australia. Saltwater mosquito production in mangroves and salt marsh is the predominant mosquito control concern in all the areas. Urban encroachment towards saltwater mosquito habitats is a problem in both Florida and Australia. In east-central Florida and the Northern Territory, mosquito control is supported by comprehensive source reduction programs, whereas in Queensland and New South Wales, larviciding is the main method of control. The long-term control by source reduction programs reduces vulnerability to mosquito issues as population encroaches towards wetlands, whereas larviciding programs have to respond repeatedly as problems arise. Problems from urban encroachment are exacerbated if mosquito control and land-use planning are not integrated. Further, urban planning that is not informed by mosquito management can lead to increased mosquito problems by inadvertent design or allowing residential development close to mosquito habitats. This increases the need for mosquito control and related resourcing. At the regional level of governance, Florida and the Northern Territory generally have greater integration between planning for development and mosquito control than at the local government level in New South Wales and Queensland, where there is a lack of integration between mosquito agencies and planners. It is concluded that coordination of planning and mosquito control is more effective at higher government levels than at local levels, which have less connectivity between management areas and/or insufficient resources. The lesson is that collaboration can assist in avoiding or resolving conflicts.

We present results from experiments at the Linac Coherent Light Source (LCLS) demonstrating that serial femtosecond crystallography (SFX) can be performed to high resolution (~2.5 Å) using protein microcrystals deposited on an ultra-thin silicon nitride membrane and embedded in a preservation medium at room temperature. Data can be acquired at a high acquisition rate using x-ray free electron laser sources to overcome radiation damage, while sample consumption is dramatically reduced compared to flowing jet methods. We achieved a peak data acquisition rate of 10 Hz with a hit rate of ~38%, indicating that a complete data set could be acquired in about one 12-hour LCLS shift using the setup described here, or in even less time using hardware optimized for fixed target SFX. This demonstration opens the door to ultra low sample consumption SFX using the technique of diffraction-before-destruction on proteins that exist in only small quantities and/or do not produce the copious quantities of microcrystals required for flowing jet methods.

X-ray diffraction patterns from two-dimensional (2-D) protein crystals obtained using femtosecond X-ray pulses from an X-ray free-electron laser (XFEL) are presented. To date, it has not been possible to acquire transmission X-ray diffraction patterns from individual 2-D protein crystals due to radiation damage. However, the intense and ultrafast pulses generated by an XFEL permit a new method of collecting diffraction data before the sample is destroyed. Utilizing a diffract-before-destroy approach at the Linac Coherent Light Source, Bragg diffraction was acquired to better than 8.5 Å resolution for two different 2-D protein crystal samples each less than 10 nm thick and maintained at room temperature. These proof-of-principle results show promise for structural analysis of both soluble and membrane proteins arranged as 2-D crystals without requiring cryogenic conditions or the formation of three-dimensional crystals.

Abomasal carnitine infusion during acute feed restriction increases hepatic fatty acid oxidation and decreases liver lipid in dairy cows. Eight mid-lactation Holstein cows were used in a replicated 4×4 Latin square design with 14-d periods. A 2×2 factorial arrangement was used to determine the effects of water infusion+ad libitum dry matter intake (DMI), water infusion+restricted DMI (50% of previous 5-d average), l-carnitine infusion (20 g/d)+ad libitum DMI, or l-carnitine infusion+restricted DMI. Liver RNA from 7 healthy cows was used for transcriptome profiling using a bovine microarray. An ANOVA with a false discovery rate was used to identify treatment and interaction effects. A substantial transcriptome change was observed only with DMI restriction, resulting in 312 (155 downregulated, 157 upregulated) differentially expressed genes. Quantitative PCR was performed to verify microarray data and measure expression of additional genes not present on the microarray. The quantitative PCR data confirmed the effect of feed restriction but not of l-carnitine treatment. Feed restriction increased expression of GPX3 and of genes associated with gluconeogenesis (PC, PDK4), inflammation (SAA3), and signaling (ADIPOR2). In contrast, feed restriction downregulated BBOX, a key for l-carnitine biosynthesis, and the transcription factor HNF4A. The bioinformatics functional analysis of genes affected by DMI restriction uncovered biosynthesis of cholesterol and energy generation by mitochondrial respiration as the most relevant and inhibited functions. The data also indicated an increase of flux toward gluconeogenesis. We interpreted those results as a likely response of the liver to spare energy and provide glucose for the lactating mammary gland during feed deprivation., (Copyright © 2013 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Here we present a novel laboratory-based cryogenic soft X-ray microscope for whole cell tomography of frozen hydrated samples. We demonstrate the capabilities of this compact cryogenic microscope by visualizing internal subcellular structures of Saccharomyces cerevisiae cells. The microscope is shown to achieve better than 50 nm half-pitch spatial resolution with a Siemens star test sample. For whole biological cells, the microscope can image specimens up to 5 μm thick. Structures as small as 90 nm can be detected in tomographic reconstructions following a low cumulative radiation dose of only 7.2 MGy. Furthermore, the design of the specimen chamber utilizes a standard sample support that permits multimodal correlative imaging of the exact same unstained yeast cell via cryo-fluorescence light microscopy, cryo-soft X-ray microscopy, and cryo-transmission electron microscopy. This completely laboratory-based cryogenic soft X-ray microscope will enable greater access to three-dimensional ultrastructure determination of biological whole cells without chemical fixation or physical sectioning.

An outstanding unresolved question is how does the mitotic spindle utilize microtubules and mitotic motors to coordinate accurate chromosome segregation during mitosis? This process depends upon the mitotic motor, kinesin-5, whose unique bipolar architecture, with pairs of motor domains lying at opposite ends of a central rod, allows it to crosslink microtubules within the mitotic spindle and to coordinate their relative sliding during spindle assembly, maintenance and elongation. The structural basis of kinesin-5's bipolarity is, however, unknown, as protein asymmetry has so far precluded its crystallization. Here we use electron microscopy of single molecules of kinesin-5 and its subfragments, combined with hydrodynamic analysis plus mass spectrometry, circular dichroism and site-directed spin label electron paramagnetic resonance spectroscopy, to show how a staggered antiparallel coiled-coil 'BASS' (bipolar assembly) domain directs the assembly of four kinesin-5 polypeptides into bipolar minifilaments.

The coupling of cryo-light microscopy (cryo-LM) and cryo-electron microscopy (cryo-EM) poses a number of advantages for understanding cellular dynamics and ultrastructure. First, cells can be imaged in a near native environment for both techniques. Second, due to the vitrification process, samples are preserved by rapid physical immobilization rather than slow chemical fixation. Third, imaging the same sample with both cryo-LM and cryo-EM provides correlation of data from a single cell, rather than a comparison of "representative samples". While these benefits are well known from prior studies, the widespread use of correlative cryo-LM and cryo-EM remains limited due to the expense and complexity of buying or building a suitable cryogenic light microscopy stage. Here we demonstrate the assembly, and use of an inexpensive cryogenic stage that can be fabricated in any lab for less than $40 with parts found at local hardware and grocery stores. This cryo-LM stage is designed for use with reflected light microscopes that are fitted with long working distance air objectives. For correlative cryo-LM and cryo-EM studies, we adapt the use of carbon coated standard 3-mm cryo-EM grids as specimen supports. After adsorbing the sample to the grid, previously established protocols for vitrifying the sample and transferring/handling the grid are followed to permit multi-technique imaging. As a result, this setup allows any laboratory with a reflected light microscope to have access to direct correlative imaging of frozen hydrated samples.

To examine effects of maternal nutrition and Se intake on adiposity and insulin sensitivity in female offspring, treatments were imposed during gestation on 82 pregnant primiparous Rambouillet ewe lambs (52.2 ± 0.8 kg) allotted randomly to 1 of 6 treatments in a 2 × 3 factorial arrangement. Factors were adequate (9.5 μg Se·kg BW(-1)·d(-1); ASe) or high (81.8 μg Se·kg BW(-1)·d(-1); HSe) levels of dietary Se (Se-enriched yeast) and maternal nutritional intake (100% of metabolizable energy [ME] requirement [MOD], 60% of MOD [LOW], and 140% of MOD [HIGH]). Selenium treatments were initiated at breeding and global nutritional treatments at day 50 of gestation. At parturition, lambs were removed from ewes before nursing and managed similarly. Glucose tolerance tests were performed at 107 and 148 d of age. Necropsies were performed at 180 d of age. Although there was no effect of Se on maternal body condition or weight during gestation, both maternal nutritional intake and selenium treatment influenced (P ≤ 0.04) offspring growth and response to a glucose tolerance test. Female lambs from HSe ewes were heavier (P = 0.04) at birth. There were nutritional intake and Se interactions (P ≤ 0.05) on the growth rate of the lambs and their insulin response to a glucose bolus at 2 different times during growth. By 180 d, ewe lambs from HSe ewes had more (P ≤ 0.07) internal fat stores than lambs from ASe ewes. It appears that both maternal nutritional level and Se intake can influence insulin sensitivity, and maternal Se intake alone can enhance fat deposition in female offspring., (Copyright © 2010 Elsevier Inc. All rights reserved.)

The objectives were to examine effects of dietary Se supplementation and nutrient restriction during defined periods of gestation on maternal adaptations to pregnancy in primigravid sheep. Sixty-four pregnant Western Whiteface ewe lambs were assigned to treatments in a 2 x 4 factorial design. Treatments were dietary Se [adequate Se (ASe; 3.05 microg/kg of BW) vs. high Se (HSe; 70.4 microg/kg of BW)] fed as Se-enriched yeast, and plane of nutrition [control (C; 100% of NRC requirements) vs. restricted (R; 60% of NRC requirements]. Selenium treatments were fed throughout gestation. Plane of nutrition treatments were applied during mid (d 50 to 90) and late gestation (d 90 to 130), which resulted in 4 distinct plane of nutrition treatments [treatment: CC (control from d 50 to 130), RC (restricted from d 50 to 90, and control d 90 to 130), CR (control from d 50 to 90, and restricted from d 90 to 130), and RR (restricted from d 50 to 130)]. All of the pregnant ewes were necropsied on d 132 +/- 0.9 of gestation (length of gestation approximately 145 d). Nutrient restriction treatments decreased ewe ADG and G:F, as a result, RC and CR ewes had similar BW and maternal BW (MBW) at necropsy, whereas RR ewes were lighter than RC and CR ewes. From d 90 to 130, the HSe-CC ewes had greater ADG (Se x nutrition; P = 0.05) than did ASe-CC ewes, whereas ADG and G:F (Se x nutrition; P = 0.08) were less for HSe-RR ewes compared with ASe-RR ewes. The CR and RR treatments decreased total gravid uterus weight (P = 0.01) as well as fetal weight (P = 0.02) compared with RC and CC. High Se decreased total (g; P = 0.09) and relative heart mass (g/kg of MBW; P = 0.10), but increased total and relative mass of liver (P < or = 0.05) and perirenal fat (P < or = 0.06) compared with ASe. Total stomach complex mass was decreased (P < 0.01) by all the nutrient restriction treatments, but was reduced to a greater extent in CR and RR compared with RC. Total small intestine mass was similar between RC and CC ewes, but was markedly reduced (P < 0.01) in CR and RR ewes. The mass of the stomach complex and the small and large intestine relative to MBW was greater (P = 0.01) for RC than for CR ewes. Increased Se decreased jejunal DNA concentration (P = 0.07), total jejunal cell number (P = 0.03), and total proliferating jejunal cell number (P = 0.05) compared with ASe. These data indicate that increased dietary Se affected whole-body and organ growth of pregnant ewes, but the results differed depending on the plane of nutrition. In addition, the timing and duration of nutrient restriction relative to stage of pregnancy affected visceral organ mass in a markedly different fashion.

This study compares mosquito control within similar environments between 26 and 30 degrees of latitude on the east central coasts of Florida and of Australia. It describes and compares the relevant mosquito-producing environments, the development of mosquito control, legislative framework, funding arrangements, and organizational differences between the areas, including the international interactions that have facilitated good practices. The article identifies some strengths and weaknesses of the programs in each area. Significant strengths include some aspects of funding and administration; collaborations with other organizations; the roles of national and state organizations, including research agencies; and commitment of individuals. Potential weaknesses in programs that are part of larger organizations include their relatively low position in the organizational hierarchy and the need to compete for resources. Programs that are independent districts may lack opportunity to interact with other land management units. Other weaknesses include the relatively high turnover of staff in state environmental resource agencies and the potential loss of institutional memory when long-term mosquito control staff members leave. The case comparison highlights similarities in product use at the individual program level (Indian River Mosquito Control District and Gold Coast Pest Management Unit) and differences in practices including aerial adulticiding being used in Florida but not in Australia.

To examine effects of nutritional plane and Se supplementation on colostrum quality and mammary development, individually fed, pregnant Rambouillet ewe lambs were allotted randomly to 1 of 6 treatments in a 2 x 3 factorial arrangement. Main effects included dietary Se level, which began at breeding (d = 0) [adequate Se (9.5 mug/kg of BW) vs. high Se (81.8 mug/kg of BW)], and plane of nutrition, which began at d 50 of gestation [60% (RES), 100% (CON), and 140% (HIGH) of requirements]. Upon parturition, lambs were immediately separated from dams and weighed. Three hours after lambing, colostrum yield was determined, and samples were obtained for components and immunoglobulin G (IgG) analysis. Ewes were slaughtered within 24 h of parturition, and mammary tissues were collected for determination of alveolar secretory epithelial cell proliferation index and luminal area. Gestation length was reduced (P < 0.01) in HIGH ewes compared with RES and CON ewes. Although birth weights were reduced (P < 0.01) in RES and HIGH compared with CON ewes, there was little effect of diet on placental size. Mammary gland weight was reduced (P /= 0.15) on mammary gland weight, colostrum quantity, or IgG concentration in pregnant ewe lambs. Improper nutrition from mid to late pregnancy in ewe lambs altered colostrum quality and quantity and reduced offspring birth weight, which may have negative implications for lamb health and survival during the early postnatal period.

The objectives of this study were to determine the effects of dietary L-carnitine supplementation on liver lipid accumulation, hepatic nutrient metabolism, and lactation in multiparous cows during the periparturient period. Cows were assigned to treatments at d -25 relative to expected calving date and remained on the experiment until 56 d in milk. Treatments were 4 amounts of supplemental dietary carnitine: control (0 g/d of L-carnitine; n = 14); low carnitine (LC, 6 g/d; n = 11); medium carnitine (MC, 50 g/d; n = 12); and high carnitine (HC, 100 g/d; n = 12). Carnitine was supplied by mixing a feed-grade carnitine supplement with 113.5 g of ground corn and 113.5 g of dried molasses, which was then fed twice daily as a topdress to achieve desired daily carnitine intakes. Carnitine supplementation began on d -14 relative to expected calving and continued until 21 d in milk. Liver and muscle carnitine concentrations were markedly increased by MC and HC treatments. Milk carnitine concentrations were elevated by all amounts of carnitine supplementation, but were greater for MC and HC than for LC during wk 2 of lactation. Dry matter intake and milk yield were decreased by the HC treatment. The MC and HC treatments increased milk fat concentration, although milk fat yield was unaffected. All carnitine treatments decreased liver total lipid and triacylglycerol accumulation on d 10 after calving. In addition, carnitine-supplemented cows had higher liver glycogen during early lactation. In general, carnitine supplementation increased in vitro palmitate beta-oxidation by liver slices, with MC and HC treatments affecting in vitro palmitate metabolism more potently than did LC. In vitro conversion of Ala to glucose by liver slices was increased by carnitine supplementation independent of dose. The concentration of nonesterified fatty acids in serum was not affected by carnitine. As a result of greater hepatic fatty acid beta-oxidation, plasma beta-hydroxybutyric acid was higher for the MC and HC treatments. Serum insulin was greater for all carnitine treatments, although plasma glucose was unaffected. Plasma urea N was lower and plasma total protein was higher for the MC and HC treatments. By decreasing liver lipid accumulation and stimulating hepatic glucose output, carnitine supplementation might improve glucose status and diminish the risk of developing metabolic disorders during early lactation.

Previously we determined that abomasal infusion of L-carnitine increased in vitro hepatic fatty acid oxidation, decreased liver lipid accumulation, and supported higher fat-corrected milk yield in feed-restricted lactating cows. The objectives of this study were to examine the effects of supplemental L-carnitine and amount of feed intake on free carnitine and carnitine ester concentrations in liver, muscle, milk, and plasma of lactating dairy cows. Eight lactating Holstein cows (132 +/- 36 d in milk) were used in a replicated 4 x 4 Latin square design with 14-d periods to test factorial combinations of water or L-carnitine infusion (20 g/d; d 5 to 14) and ad libitum or restricted (50% of previous 5-d intake; d 10 to 14) dry matter intake. Plasma was obtained 3 times daily on d 4, 8, and 12; milk samples were collected on d 8, 9, 13, and 14. Liver and muscle were biopsied on d 14 of each period. Free carnitine, short-chain acylcarnitine, and long-chain acylcarnitine concentrations were determined using a radioenzymatic assay coupled with ion exchange chromatography. Abomasal L-carnitine infusion increased total carnitine in plasma on d 8 and d 12. All liver carnitine fractions were increased by carnitine infusion. Feed restriction elevated concentrations of free carnitine, long-chain acylcarnitine, and total carnitine in liver tissue from carnitine-infused cows but not in those infused with water. In muscle, acid-soluble carnitine, long-chain acylcarnitine, and total carnitine concentrations were increased by carnitine infusion and feed restriction without significant interaction. Feed restriction increased free carnitine concentrations in muscle from water-infused cows but not in carnitine-infused cows. Carnitine infusion increased the concentration of each milk carnitine fraction as well as milk carnitine output on d 8 to 9. On d 13 to 14, all carnitine fractions except short-chain acylcarnitine were increased in milk from water-infused, feed-restricted cows, whereas all fractions were increased in carnitine-infused, feed-restricted cows. Carnitine infusion increased total carnitine in plasma, liver, muscle, and milk during feed restriction, whereas feed restriction alone increased carnitine concentrations in muscle and milk but not in liver. Liver carnitine concentrations might limit hepatic fatty acid oxidation capacity in dairy cows during the periparturient period; therefore, supplemental L-carnitine might decrease liver lipid accumulation in periparturient cows.

Selection of appropriate housekeeping genes (HKG) for normalization of quantitative PCR data for genes of interest is critical for interpretation of results. Ideally, copy number of the chosen HKG mRNA will not vary with experimental treatments or physiological state in the tissue studied, which improves accuracy in detecting changes in genes of interest. Because of the liver's dynamic role in metabolism, physiological state or dietary treatments could alter mRNA expression of commonly used HKG. Therefore, the objective of this study was to evaluate stability of mRNA expression for a number of candidate HKG in bovine liver across different physiological and dietary experimental conditions during the periparturient period. A publicly available program (geNorm) was used to evaluate expression stability of 8 HKG (beta-actin, glyceraldehyde 3-phosphate dehydrogenase, beta-glucuronidase, peptidylprolyl isomerase A, polyubiquitin, ribosomal protein S9, ribosomal protein L32, and 18S ribosomal RNA) in 91 liver RNA samples. Screened samples included liver from cows in 3 groups: 1) cows receiving a dietary supplement pre- and postpartum (n = 10); 2) cows with clinical or subclinical ketosis (n = 7); and 3) cows consuming different amounts of energy prepartum (n = 74). In group 3, samples from d -65, -30, -14, 1, 14, 28, and 49 relative to parturition were included to enable characterization of HKG mRNA expression across different physiological states. Initial analyses indicated that mRNA for ribosomal protein S9 (RPS9) was one of the most stably expressed across different experiment types. To determine the best gene, 200 bootstrap replications of the original data set were performed to determine if the ranking of RPS9 was superior to the other 7 genes evaluated. Average ranks and estimated standard errors for the top 3 genes were 1.64 +/- 0.06, 3.27 +/- 0.10, and 3.71 +/- 0.12 for RPS9, GAPDH, and beta-actin, respectively. Ribosomal protein S9 was ranked first 59% of the time and was never ranked lower than fifth. The lowest-ranked gene was polyubiquitin, ranked last 46.5% of the time (average rank = 6.85 +/- 0.10). In this study, physiological state, amount of intake, or dietary treatment influenced the mRNA expression of commonly used HKG in bovine liver. Ideally, expression stability should be tested before collection of data in all experiments; however, we have shown that RPS9 mRNA is stable across several physiological and diet-related experimental conditions for dairy cows, making it a good HKG in liver quantitative PCR experiments.

L-Carnitine is required for mitochondrial fatty acid oxidation, but the effects of carnitine supplementation on nutrient metabolism during dry matter intake depression have not been determined in dairy cows. Studies in other species have revealed responses to L-carnitine that may be of specific benefit to dairy cows during the periparturient period. Eight lactating Holstein cows (132 +/- 36 d in milk) were used in a replicated 4 x 4 Latin square experiment with 14-d periods. Treatments were factorial combinations of abomasal infusion of either water or L-carnitine (20 g/d; d 5 to 14) and either ad libitum or restricted intake (50% of previous 5-d dry matter intake; d 10 to 14) of a balanced lactation diet. Liver and muscle biopsies were obtained on d 14 of each period. Feed restriction induced negative balances of energy and metabolizable protein. In feed-restricted cows, carnitine infusion increased 3.5% fat-corrected milk yield compared with those infused with water. Total carnitine concentration in liver was increased in feed-restricted cows infused with carnitine but not in feed-restricted cows infused with water. Carnitine infusion stimulated in vitro oxidation of [1-(14)C] palmitate to acid-soluble products and decreased the proportion of [1-(14)C] palmitate that was converted to esterified products by liver slices. Feed-restricted cows infused with carnitine had lower liver total lipid concentration and tended to have decreased triglyceride accumulation compared with feed-restricted cows infused with water. Plasma nonesterified fatty acid concentration was not altered by carnitine infusion but was increased by feed restriction; serum beta-hydroxybutyric acid was increased by carnitine infusion in feed-restricted cows. In cows fed for ad libitum intake, carnitine infusion affected beta-hydroxybutyric acid, insulin, and urea N in serum, liver glycogen concentration, and in vitro alanine oxidation by liver slices, suggesting that hepatic and peripheral nutrient metabolism was influenced. L-Carnitine infusion effectively decreased liver lipid accumulation during feed restriction as a result of greater capacity for hepatic fatty acid oxidation. Further research examining dietary supplementation of L-carnitine during the periparturient period is warranted.

Source reduction as part of an integrated pest management program is a cornerstone of the American Mosquito Control Association's Pesticide Environmental Stewardship Program Strategy Document to reduce pesticide risk. Since the early 1980s, Florida has made important strides in implementing environmentally sound source reduction strategies in salt marshes while managing them for both mosquito control and natural resource enhancement. The political mechanism for this progress has been interagency cooperation through the Florida Coordinating Council on Mosquito Control and its Subcommittee on Managed Marshes. Challenges in accomplishing source reduction continue because both public and private lands are involved. Public lands include those owned by federal (e.g., U.S. Fish and Wildlife Service, National Park Service), state (Florida Department of Environmental Protection), and local governments, and they have a diversity of management objectives. This diversity adds to the challenge facing mosquito control agencies in providing mosquito control services while protecting and enhancing the environment.

Choline is important for assembly of very low density lipoproteins to export triglyceride from liver; however, studies to assess the effect of rumen-protected choline (RPC) supplementation on blood lipid metabolites in periparturient dairy cows have not been conducted. Thirty-two multiparous Holstein and 10 multiparous Jersey cows were randomly assigned to control or RPC treatments. A close-up diet was fed from approximately 3 wk before parturition through parturition, followed by a lactation diet from parturition through 49 d postpartum. For RPC, diets were top-dressed once daily with 60 g of a RPC product (25% choline as choline chloride) from 21 d before expected parturition through 21 d postpartum. Treatment did not affect dry matter intake either prepartum (12.0 vs. 12.1 kg/d for RPC and control, respectively) or during the first 3 wk postpartum (14.8 vs. 15.7 kg/d, respectively). Daily yields of 3.5% fat-corrected milk (39.4 vs. 37.4 kg/d), fat (1.46 vs. 1.38 kg/d), and protein (1.09 vs. 1.05 kg/d) did not differ statistically by treatment (RPC vs. control, respectively). Jersey cows in the control group had lower concentrations of nonesterified fatty acids and beta-hydroxybutyrate in plasma during d 1 to 10 postpartum than did other breed and treatment combinations. Cows fed RPC tended to have greater serum triglycerides prepartum (17.0 vs. 14.7 mg/dL) and lower plasma phospholipid at parturition (65.2 vs. 78.1 mg/dL) than control cows. Treatment did not affect cholesterol and phospholipid at other time points, but concentrations followed patterns of dry matter intake pre- and postpartum. Cows were in moderate body condition score (mean = 3.3) at the start of the study and did not lose excessive condition by 3 wk postpartum (mean body condition score loss = 0.5); therefore, cows might not have been at great risk for hepatic lipid accumulation. Additionally, calculated Met balance was negative postpartum; supplemental RPC might not have spared enough Met to produce a physiological benefit. More research is needed to determine how choline affects prevention or alleviation of fatty liver syndrome and to confirm potential differences between Holstein and Jersey cows.

The cytosolic Ah receptor (AhR) heterocomplex consists of one molecule of the AhR, a 90-kDa heat shock protein (Hsp90) dimer, and one molecule of the hepatitis B virus X-associated protein 2 (XAP2). Serine residues 43,53,131-2, and 329 on XAP2-FLAG were identified as putative phosphorylation sites using site-directed mutagenesis followed by two-dimensional phosphopeptide mapping analysis. Protein kinase CK2 (CK2) was identified as the 45-kDa kinase from COS 1 cell or liver extracts that was responsible for phosphorylation of serine 43 in the XAP2 peptide 39-57. Loss of phosphorylation at any or all of the serine residues did not significantly affect the ability of XAP2-FLAG to bind to the murine AhR in rabbit reticulocyte lysate or Hsp90 in COS-1 cells. Furthermore, all of these serine mutants were able to sequester murine AhR-YFP into the cytoplasm as well as wild-type XAP2. YFP-XAP2 S53A was unable to enter the nucleus, indicating a potential role of phosphorylation in nuclear translocation of XAP2.

The aryl hydrocarbon (Ah) receptor (AhR) is a member of the basic helix-loop-helix PER-ARNT-SIM (PAS) transcription factor family. Consistent with the notion that PAS proteins are biological sensors, AhR binding to Ah toxicants induces or represses transcription of a wide range of genes and results in a cascade of toxic responses. However, an endogenous role for AhR in development and homeostasis is supported by (1) the discovery of low affinity, endogenous ligands; (2) studies demonstrating a role for the receptor in development of liver and vascular systems, that were established using mice lacking AhR expression; and (3) the presence of functional dioxin-responsive elements in promoter regions of genes involved in cellular growth and differentiation. A large body of recent literature has implicated AhR in multiple signal transduction pathways. AhR is known to interact with signaling pathways that are mediated by estrogen receptor and other hormone receptors, hypoxia, nuclear factor kappaB, and retinoblastoma protein. In addition, AhR complexes may affect cellular signaling through interactions with various other regulatory and signaling proteins, including PAS heterodimerization partners (ARNT), chaperone and immunophilin-like proteins (e.g. HSP90, XAP2/ARA9/AIP, p23), protein kinases and phosphatases (e.g. tyrosine kinases, casein kinase 2, protein kinase C), and coactivators (e.g. SRC-1, RIP 140, CBP/p300). Here we summarize the types of molecular cross talk that have been identified between AhR and cell signaling pathways., (Copyright 2002 Wiley Periodicals, Inc. J Biochem Mol Toxicol 16:317-325, 2002; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.10051)

There is a great deal of evidence that altered sphingolipid metabolism is associated with fumonisin-induced animal diseases including increased apoptotic and oncotic necrosis, and carcinogenesis in rodent liver and kidney. The biochemical consequences of fumonisin disruption of sphingolipid metabolism most likely to alter cell regulation are increased free sphingoid bases and their 1-phosphates, alterations in complex sphingolipids, and decreased ceramide (CER) biosynthesis. Because free sphingoid bases and CER can induce cell death, the fumonisin inhibition of CER synthase can inhibit cell death induced by CER but promote free sphingoid base-induced cell death. Theoretically, at any time the balance between the intracellular concentration of effectors that protect cells from apoptosis (decreased CER, increased sphingosine 1-phosphate) and those that induce apoptosis (increased CER, free sphingoid bases, altered fatty acids) will determine the cellular response. Because the balance between the rates of apoptosis and proliferation is important in tumorigenesis, cells sensitive to the proliferative effect of decreased CER and increased sphingosine 1-phosphate may be selected to survive and proliferate when free sphingoid base concentration is not growth inhibitory. Conversely, when the increase in free sphingoid bases exceeds a cell's ability to convert sphinganine/sphingosine to dihydroceramide/CER or their sphingoid base 1-phosphate, then free sphingoid bases will accumulate. In this case cells that are sensitive to sphingoid base-induced growth arrest will die and insensitive cells will survive. If the cells selected to die are normal phenotypes and the cells selected to survive are abnormal, then cancer risk will increase.

Laboratory studies have described the carcinogenicity of fumonisin B1 (FB1) in rodents and epidemiological evidence suggests an association between FB1 (a mycotoxin produced by Fusarium moniliforme) and cancer in humans. This study was designed to reveal in rainbow trout, a species with very low spontaneous tumor incidence, if FB1 was (i) a complete carcinogen, in the absence of an initiator; (ii) a promoter of liver tumors in fish initiated as fry with aflatoxin B1 (AFB1); and (iii) a promoter of liver, kidney, stomach, or swim bladder tumors in fish initiated as fry with N-methyl-N'-nitro-nitrosoguanidine (MNNG). FB1 was not a complete carcinogen in trout. No tumors were observed in any tissue of fish fed diets containing 0, 3.2, 23, or 104 ppm FB1 for a total of 34 weeks (4 weeks FB1 exposure, 2 weeks outgrowth on control diet, followed by 30 weeks FB1 diet) in the absence of a known initiator. FB1 promoted AFB1 initiated liver tumors in fish fed > or = 23 ppm FB1 for 42 weeks. A 1-week pretreatment of FB1 did not alter the amount of liver [3H]AFB1 DNA adducts, which suggests that short-term exposure to FB1 will not alter phase I or phase II metabolism of AFB1. In MNNG-initiated fish, liver tumors were promoted in the 104 ppm FB1 treatment (42 weeks), but FB1 did not promote tumors in any other tissue. Tumor incidence decreased in kidney and stomach in the 104 ppm FB1 treatment of MNNG-initiated trout. The FB1 promotional activity in AFB1-initiated fish was correlated with disruption of sphingolipid metabolism, suggesting that alterations in associated sphingolipid signaling pathways are potentially responsible for the promotional activity of FB1 in AFB1-initiated fish., (Copyright 2001 Academic Press.)

Many natural and synthetic xenobiotics are known to interact with endocrine systems of animals. Various hydroxylated metabolites of persistent polychlorinated biphenyl contaminants (hydroxy-polychlorinated biphenyls [OH-PCBs]) have been shown to have agonist or antagonist interactions with estrogen receptors (ERs). In this study, 4-hydroxy-2',4',6'-trichlorobiphenyl (OH-PCB 30) and 4-hydroxy-2',3',4',5'-tetrachlorobiphenyl (OH-PCB 61), and the natural estrogen 17 beta-estradiol (E2) and estrone (E1), were incorporated into diet and fed to juvenile rainbow trout. The production of vitellogenin (VTG), an egg yolk protein precursor in oviparous animals, was used as a marker of hepatic ER binding. All compounds induced plasma VTG in a dose-dependent manner, with maximal levels of approximately 5 mg VTG/ml plasma induced by E2, E1, and OH-PCB 30. Maximum plasma VTG of 0.048 mg/ml in the highest dose (50 mg/kg) of OH-PCB 61 was approximately 100-fold lower than natural estrogens and OH-PCB 30. At doses that induced submaximal VTG, E1 was two- to threefold less potent, and OH-PCBs were up to 500-fold less potent, than E2. Sex differences in VTG synthesis were apparent at weakly estrogenic doses, but not at maximal VTG-inducing doses. Predictions from previous receptor-binding studies underestimated the maximum estrogenic response of OH-PCB 30 in trout, which was achieved with a dose 10 times higher than E2. Differences in plasma VTG induction by OH-PCB 30 and OH-PCB 61 support in vitro predictions that the degree and position of chlorination are important for ER activation. Neither mixtures of estrogens nor OH-PCBs resulted in synergistic VTG induction.

Indole-3-carbinol (I3C), a compound found in Brassica vegetables has been widely studied for its chemopreventive properties. I3C has been shown to block tumor initiation and promotion; however, it also acts as a tumor promoter. I3C and some of its acid condensation products, particularly 3,3'-diindolylmethane (I33'), have exhibited antiestrogenic properties. We report that I33' acts as an estrogen in the rainbow trout liver in vitro and in vivo by inducing vitellogenin (Vg), a well-characterized biomarker for estrogens. Precision-cut liver slices from male rainbow trout, Oncorhynchus mykiss, were incubated at 14 degrees C for 96 h in media containing I3C, I33', or a mixture of I3C acid condensation products (RXN) (0-250 microM). I33' and RXN increased Vg levels in rainbow trout liver slices by over 300- and 20-fold, respectively, vs vehicle. The efficacy of I33' induction of Vg was comparable to 17 beta-estradiol (E(2)) with 2500-fold less potency. I33' and E(2) cotreatment resulted in additive Vg induction. Tamoxifen completely inhibited I33'-induced Vg induction, suggesting that Vg induction by I33' is entirely through the estrogen receptor. In vivo, juvenile male rainbow trout were fed I3C, RXN (0-2000 mg/kg), or I33' (0-250 mg/kg) for 2 weeks. At 2000 mg/kg, I3C induced Vg by over 100,000-fold compared to controls, which was comparable to 5 mg/kg 17 beta-estradiol (the dose resulting in maximum induction). I33' was five times as potent as I3C with equal efficacy. The potency of RXN was only 5% of I3C. Again, I33' and E(2) cotreatment resulted in additive Vg induction. I33' may have accounted for Vg increases observed in trout fed I3C as it is present in liver after oral dosing at concentrations (70 microM) expected to maximally induce Vg. In trout, results in vitro and in vivo document that I33' is estrogenic, consistent with our hypothesis that I3C promotes liver cancer in trout by estrogenic pathways., (Copyright 2001 Academic Press.)

Fish sexual development is sensitive to exogenous hormone manipulation, and salmonids have been used extensively as environmental sentinels and models for biomedical research. We simulated maternal transfer of contaminants by microinjecting rainbow trout (Oncorhynchus mykiss) and chinook salmon (Oncorhynchus tshawytscha) embryos. Fish were reared for 6 months and sexed, and gonads were removed for histology and measurement of in vitro steroid production. Analysis of fat samples showed that dichlorodiphenylethylene (DDE) levels, o, p'M-DDE and p,o, p'-DDE isomers, were elevated 6 months after treatment. A preliminary study showed an increased ratio of males to females after treatment with 80 mg/kg and 160 mg/kg of the xenoestrogen o,o, p'-DDE. One fish treated with 160 mg/kg o,o, p'-DDE had gonads with cells typical of both males and females. A follow-up study, using more fish and excluding the highly toxic 160 mg/kg o,o, p'-DDE dose, showed no effect on sex ratio or gonadal histology. Embryonic exposure of monosex male trout, monosex female trout, and mixed sex salmon to o, o, p'-DDE, p,o, p'-DDE, mixtures of DDE isomers, and octylphenol failed to alter sexual development. We observed no treatment-dependent changes in in vitro gonadal steroid production in any experiments. Trout exposed in ovo and reared to maturity spawned successfully. These results suggest that mortality attributable to the xenoestrogens o,o, p'-DDE, chlordecone, and octylphenol, and the antiandrogen p,o, p'-DDE, is likely to occur before the appearance of subtle changes in sexual development. Because trout appeared to be sensitive to endocrine disruption, we cannot dismiss the threat of heavily contaminated sites or complex mixtures to normal sexual development of salmonids or other aquatic organisms.

Vitellogenin (Vg) was measured in sexually immature rainbow trout (Oncorhynchus mykiss) exposed to 17β-estradiol (E 2 ) in the diet. Mixed-sex populations of trout aged 3, 6, 12, or 18 months were maintained separately and fed E 2 at 0.05 or 2.5 mg/kg for 7 d. Females fed E 2 at 0.05 mg/kg consistently produced three- to fourfold greater amounts of Vg than similarly aged males. Age- and sex-matched fish fed E 2 at 2.5 mg/kg produced equivalent amounts of Vg. Sex differences in Vg production were apparent only at a dose of E 2 (0.05 mg/kg) that results in submaximal Vg induction. Our results document the importance of considering the sex of juvenile fish when using Vg production as a marker of xenoestrogen exposure., (Copyright © 1999 SETAC.)

The feasibility of utilizing rainbow trout, Oncorhynchus mykiss, as an alternative model for studying the inhibition of aromatase (CYP 19) was investigated. The suppression of estrogen-dependent tumors by aromatase inhibitors has been important in the treatment of breast cancer. Estrogens, estrogen precursors and xenoestrogens have been found to promote liver cancer in the trout model. A steroid, 4-hydroxy-4-androstene-3,17-dione (4-OHA), and non-steroids, aminoglutethimide (AG) and Letrozole (CGS 20267), all of which are known aromatase inhibitors in rats and humans, were examined in vitro for activity in trout ovarian microsomes. Aromatase activity was quantified as the release of 3H2O from the conversion of [3H]-4-androstene-3,17-dione to 17beta-estradiol and estrone. Trout ovarian microsomes exhibited activity between 39-60 fmol mg(-1) min(-1) with a calculated Vmax of 71.1 fmol mg(-1) min(-1) when incubated at 25 degrees C with 200 nM 4-androstene-3,17-dione (K(M) = 435 nM). Significant inhibition by 4-OHA up to 80% was seen at 1.5 microM. At 2000 microM, AG decreased aromatase activity by up to 82%. Letrozole reduced aromatase activity a maximum of 90% in a dose-dependent manner, but the Ki (2.3 microM) was 1000-fold higher than reported in human trials. Indole-3-carbinol and some of its derivatives, two DDE isomers and four flavones (except alpha-naphthoflavone) at 1000 microM did not significantly inhibit aromatase in vitro. Letrozole and clotrimazole, fed to juvenile rainbow trout at doses up to 1000 ppm for 2 weeks, were not effective in suppressing dehydroepiandrosterone (DHEA) induced increases in vitellogenin and 17beta-estradiol levels. These results document that trout aromatase is sensitive to inhibition in vitro by known inhibitors of the mammalian enzyme. The mechanism(s) for lack of inhibition in vivo is currently unknown and must be further investigated in order to develop a trout model for studying the role of aromatase in carcinogenesis.

During the 1990s, Florida has continued to make important strides in managing salt marshes for both mosquito control and natural resource enhancement. The political mechanism for this progress continues to be interagency cooperation through the Florida Coordinating Council on Mosquito Control and its Subcommittee on Managed Marshes (SOMM). Continuing management experience and research has helped refine the most environmentally acceptable source reduction methods, which typically are Rotational Impoundment Management or Open Marsh Water Management. The development of regional marsh management plans for salt marshes within the Indian River Lagoon by the SOMM has helped direct the implementation of the best management practices for these marshes. Controversy occasionally occurs concerning what management technique is most appropriate for individual marshes. The most common disagreement is over the benefits of maintaining an impoundment in an "open" vs. "closed" condition, with the "closed" condition, allowing for summer mosquito control flooding or winter waterfowl management. New federal initiatives influencing salt-marsh management have included the Indian River Lagoon-National Estuary Program and the Pesticide Environmental Stewardship Program. A new Florida initiative is the Florida Department of Environmental Protection's Eco-system Management Program with continuing involvement by the Surface Water Improvement and Management program. A developing mitigation banking program has the potential to benefit marsh management but mosquito control interests may suffer if not handled properly. Larvicides remain as an important salt-marsh integrated pest management tool with the greatest acreage being treated with temephos, followed by Bacillus thuringiensis israelensis and methoprene. However, over the past 14 years, use of biorational larvicides has increased greatly.

A technical powder of Bacillus thuringiensis serovar. israelensis (B.t.i.) (VectoBac TP, 5,000 international toxic units [ITU]/mg), an aqueous suspension (VectoBac 12AS, 1,200 ITU/mg), and a granular formulation (VectoBac CG, 200 ITU/mg) were tested in the laboratory under different biotic and abiotic, conditions for efficacy against larvae of saltwater (Aedes taeniorhynchus) and freshwater (Culex nigripalpus) mosquitoes. Second-, 3rd-, and 4th-instar larvae of Cx. nigripalpus were 1.3-3-fold more susceptible to both VectoBac TP and VectoBac 12AS than were the respective larval instars of Ae. taeniorhynchus. For each species, 2nd-instar larvae were several-fold more susceptible to these B.t.i. preparations than were the 4th instars. In test cups, larvae under lower densities exposed to B.t.i. concentrations sustained 5-9-fold higher mortalities than larvae under high-density conditions. VectoBac TP and VectoBac 12AS stayed in suspension for up to 24 h with similar larvicidal efficacy, which was greater at 32-35 degrees C than at 15-20 degrees C. At 60 degrees C maintained for 24 h, VectoBac 12AS was adversely affected 2-3-fold in terms of potency, but VectoBac TP was not affected. Significant loss of potency of both VectoBac 12AS and VectoBac TP occurred when exposed to 35-37 degrees C under high light intensity (140,000-170,000 lux) for 6 h. Increasing salinity levels from 0 (fresh water) to 50% sea water caused gradual efficacy declines of VectoBac 12AS and VectoBac TP against Ae. taeniorhynchus larvae. VectoBac CG caused insignificant initial and residual (up to 8 days) larval mortalities of both mosquito species. This formulation did not release the active ingredient of B.t.i. in any significant mosquito larvicidal concentration in surface layers of water, and the formulation was more effective in shallower water. Storage of all 3 formulations under constant laboratory and variable field conditions for up to 8 months did not produce detectable potency loss of these products.

The purpose of this study was (i) to determine whether pure fumonisin B1 could be incorporated into, recovered, and detected by high-pressure liquid chromatographic analysis from the semipurified Oregon test diet (OTD) used in rainbow trout feeding studies, and (ii) to determine if the incorporated fumonisin B1 was biologically available using the change in free sphingoid bases in liver, kidney, and serum as a mechanism-based biomarker. The results indicate that fumonisin is not easily quantified in the OTD. Recoveries ranged from 12 to 81% of the calculated concentrations based on the fumonisin B1 added to the OTD. However, the fumonisin B1 in the OTD was readily absorbed and biologically active as evidenced by marked increases in free sphinganine in liver, kidney, and serum. The magnitude of the increase in free sphinganine at 100 ppm in the OTD was comparable to that known to be associated with liver toxicity in rats, pigs, and ponies.

For the past decade, salt marsh management in Florida has been a central issue in attempts to reconcile mosquito control and natural resource interests. Progress has been made in trying to maintain effective mosquito control while protecting and enhancing salt marsh resources primarily due to: 1) efforts by the Florida Coordinating Council on Mosquito Control and its Subcommittee on Managed Marshes, which are committees comprised of agencies responsible for wetlands resources, those mandated to provide mosquito control, and research institutions; and 2) funding of research to investigate ecosystem effects of marsh management techniques. Research and management experience have demonstrated that Rotational Impoundment Management (RIM) and rotary ditching can provide ecologically sound source reduction benefits. Salt marsh ownership, management of state lands and mariculture remain controversial salt marsh management issues.

Tests on the effect of hand applied naled thermal fog, both as a single treatment on one day/week and a single treatment on 3 successive days, did not control Wyeomyia vanduzeei and Wy. mitchellii. Five-min landing/biting counts in a native oak/palm woodland demonstrated that single applications produced an average landing rate decrease of 13%. Treatments 3 days in succession did not suppress the landing rate.

In Indian River County, Florida mosquito control impoundments, larval mosquito sampling and hydrological measurements demonstrated the importance of careful consideration of these factors when developing management plans for impoundments used for wastewater retention. Discharging secondarily treated wastewater into an impoundment resulted in only minor mosquito production. However, a treatment plant failure produced extremely high Culex densities in the impoundment. Average water loss rates in impoundments studied were due to evapotranspiration (0.25 cm/day) and percolation (0.38 cm/day). Greatest percolation (0.68 cm/day) was measured when the impoundments were maximally flooded. Under the conditions of this study, the impoundments can assimilate approximately 124 cm/year of wastewater (1.52 million liters/day) over a 50 ha area without overflows.

Seasonal and habitat influences on the egg-laying activity of four species of Culex were compared in south Florida using jar- and vat-type oviposition traps. Egg rafts of Culex nigripalpus Theobald, Cx. quinquefasciatus Say, and Cx. salinarius Coquillett were collected in every month of the year, but rafts of Cx. restuans Theobald were absent during spring and summer. Cx. nigripalpus was the dominant species in the summer and fall, and Cx. quinquefasciatus and Cx. salinarius peaked during the winter and spring. Cx. restuans oviposited most rafts in the fall (98.4%). Most Cx. nigripalpus egg rafts were laid in vats (88.5%), and most Cx. quinquefasciatus egg rafts were deposited in jars (58.7%). Fewer Cx. quinquefasciatus rafts were found in a wooded area (32.4%) with a dense undergrowth than in a more open area (67.6%), but Cx. nigripalpus showed no ovipositional preference between these two habitats. The number of rafts oviposited by all species was reduced during periods of rainfall.

In Indian River County, Florida, mosquito populations were monitored weekly for 12 months in early and more advanced treatment stages of secondarily treated domestic wastewater and also in untreated primary and secondary citrus packing house washwater. Well defined differences in mosquito species abundances for early vs. late wastewater stages existed. In the early levels of secondarily treated effluent and in the primary citrus washwater, Culex quinquefasciatus populations were dominant. In the later wastewater stages of both systems, Cx. nigripalpus, Anopheles spp. and Uranotaenia spp. were more common. Seasonal Culex spp. abundance patterns were apparent at some but not all study sites. Water chemistry measurements demonstrated that both systems contained relatively low levels of nutrient-related parameters with large concentration variability.

Over two years, the management regimes of: 1) opening a southeast Florida salt marsh impoundment to the adjacent estuary with culverts through the dike, then, 2) passively retaining water with flapgate risers was studied to determine the effects on marsh flooding and resultant mosquito production. Larval dipping demonstrated that all broods occurred at elevations of 0.25-0.90 ft (= 0.08-0.27 m) NGVD. Mosquito production differed significantly between some sampling quadrats and 65 (out of 75) broods were produced in the spring and summer from rainfall. Without artificial pumping, trapping of rainfall with flapgate risers aided in eliminating oviposition sites but still allowed mosquito production in some marsh locations. Even though tidal flooding permitted larvivorous fish access to mosquito larvae, they were not able to provide adequate control to eliminate larviciding.

Culex nigripalpus adults were collected from sentinel chickens at 4 sites in Indian River Co. (IRC), Florida. Chickens located in well drained, open habitats attracted fewer Cx. nigripalpus than did those located in a moist, forested area, but the number of mosquitoes collected in the open habitats increased significantly with heavy autumn rainfall. All St. Louis encephalitis virus (SLE) seroconversions (11/674) during a 7-year period were from chickens located in open, well drained habitats. A comparison of monthly rainfall during years of known SLE activity in IRC with the 22-year average showed that virus activity was significantly associated with years in which rainfall was unusually low in either September or October. Sentinel chickens in areas with large vector populations may not be the most reliable indicators of local SLE activity and autumn rainfall in September and October may influence yearly SLE patterns in southern Florida.

Mosquito production was monitored by larval dipping for 12 months in a 20.2 ha central east coast Florida salt marsh impoundment which was being managed under a rotational impoundment management (RIM) regime. This regime, implemented to provide mosquito control while retaining natural resource benefits, virtually eliminated salt-marsh Aedes mosquito production from late May through September when the marsh was closed to the estuary and flooded to approximately 1.0 ft NGVD. Anopheles spp. were collected only along the upland marsh edges in relatively low densities. Compared with the management methods of: 1) open to the estuary with culverts and, 2) passive retention of water with flapgate risers, RIM proved to be significantly more effective in reducing mosquito production.