Selection of suitable reference genes for normalization of RT-qPCR in three tissues of Northern bobwhite (Colinus virginianus) infected with eyeworm (Oxyspirura petrowi).

BACKGROUND: The Northern bobwhite (Colinus virginianus) is an economically important, and popular game bird in North America. Northern bobwhites have experiencing declines of > 3.5% annually in recent decades due to several factors. The eyeworm Oxyspirura petrowi is a nematode parasite frequently found in the eyes of bobwhites. Although reported frequently in wild bobwhites, there is no research to understand the host-parasite mechanism. Hence, it is important to investigate mechanisms of eyeworm invasion and immune modulation in bobwhite. Cytokine gene expression using RT-PCR is widely used to identify the innate immune response of a host to an infection. METHODOLOGY: In this study, we evaluated ten reference genes (HMBS, RPL19, RPL32, RPS7, RPS8, TATA, SDHA, YWHAZ, GAPDH, and ACTB) for their stability across three tissues (liver, spleen, and caecal tonsils) of control and O. petrowi infected Northern bobwhites. Primer efficiency and reference genes stability were assessed using GeNorm, NormFinder, and BestKeeper. RESULTS: Expression of these reference genes with respect to O. petrowi infection in bobwhites showed RPL32 and HMBS were the most stable genes in the liver, HMBS and SDHA were the most stable genes in the spleen, and HMBS and YWHAZ were equally stable reference genes in the caecal tonsils. CONCLUSION: Based on the geometric mean of all three analyses, our results indicate that the combination of RPL32 and HMBS for the liver, HMBS and SDHA for the spleen, and YWHAZ and HMBS for caecal tonsils might be used as reference genes for normalization in gene expression investigations on Northern bobwhites.

Validation of an LC-MS/MS method for assessment of fenbendazole sulfone drug residue in northern bobwhite liver.

The Northern Bobwhite (Colinus virginianus) is an economically important game bird within the Rolling Plains Ecoregion. Within this region, bobwhite is experiencing extreme cyclic population fluctuations which are resulting in a net decline in total population. It is suspected that within this region two helminth parasites, an eyeworm (Oxyspirura petrowi) and a cecal worm (Aulonocephalus pennula), are contributing to this phenomenon. However, this has been difficult to study as the primary mode of investigation would be the deployment of anthelmintic treatment. Unfortunately, no registered treatments for wild bobwhite currently exist. Thus, utilizing an anthelmintictreatment for wild bobwhite would require registration of that treatment with the U.S. Food and Drug Administration (FDA). As bobwhite are game birds that are hunted, they are considered food-producing animals to the FDA, and as such require the assessment for the withdrawal of the drug residues to be assessed for human food safety. In this study, we optimized and validated a bioanalytical method for the quantification of fenbendazole sulfone in bobwhite following the U.S. FDA Center for Veterinary Medicine Guidance for Industry #208 [VICH GL 49 (R)] for assessment of fenbendazole sulfone drug residue in Northern bobwhite liver. The official method for quantifying fenbendazole sulfone in domestic chicken (Gallus gallus) was adapted for use in bobwhite. The validated method quantitation range is 2.5-30 ng/mL for fenbendazole with an average recovery of 89.9% in bobwhite liver.

Molecular phylogenetic and in silico analysis of glyceraldeyde-3-phosphate dehydrogenase (GAPDH) gene from northern bobwhite quail (Colinus virginianus).

Many recent studies have been focused on prevalence and impact of two helminth parasites, eyeworm Oxyspirura petrowi and caecal worm Aulonocephalus pennula, in the northern bobwhite quail (Colinus virginianus). However, few studies have attempted to examine the effect of these parasites on the bobwhite immune system. This is likely due to the lack of proper reference genes for relative gene expression studies. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) is a glycolytic enzyme that is often utilized as a reference gene, and in this preliminary study, we evaluated the similarity of bobwhite GAPDH to GAPDH in other avian species to evaluate its potential as a reference gene in bobwhite. GAPDH was identified in the bobwhite full genome sequence and multiple sets of PCR primers were designed to generate overlapping PCR products. These products were then sequenced and then aligned to generate the sequence for the full-length open reading frame (ORF) of bobwhite GAPDH. Utilizing this sequence, phylogenetic analyses and comparative analysis of the exon-intron pattern were conducted that revealed high similarity of GAPDH encoding sequences among bobwhite and other Galliformes. Additionally, This ORF sequence was also used to predict the encoded protein and its three-dimensional structure which like the phylogenetic analyses reveal that bobwhite GAPDH is similar to GAPDH in other Galliformes. Finally, GAPDH qPCR primers were designed, standardized, and tested with bobwhite both uninfected and infected with O. petrowi, and this preliminary test showed no statistical difference in expression of GAPDH between the two groups. These analyses are the first to investigate GAPDH in bobwhite. These efforts in phylogeny, sequence analysis, and protein structure suggest that there is > 97% conservation of GADPH among Galliformes. Furthermore, the results of these in silico tests and the preliminary qPCR indicate that GAPDH is a prospective candidate for use in gene expression analyses in bobwhite.

Prevalence of monarch (Danaus plexippus) and queen (Danaus gilippus) butterflies in West Texas during the fall of 2018.

BACKGROUND: The monarch butterfly (Danaus plexippus) is a conspicuous insect that has experienced a drastic population decline over the past two decades. While there are several factors contributing to dwindling monarch populations, habitat loss is considered the most significant threat to monarchs. In the United States, loss of milkweed, particularly in the Midwest, has greatly reduced the available breeding habitat of monarchs. This has led to extensive efforts to conserve and restore milkweed resources throughout the Midwest. Recently, these research and conservation efforts have been expanded to include other important areas along the monarch's migratory path. RESULTS: During the fall of 2018, we conducted surveys of monarch eggs and larvae through West Texas. We documented monarch and queen butterfly (Danaus gilippus) reproduction throughout the region and used the proportion of monarch and queen larva to estimate the number of monarch eggs. Peak egg densities for monarchs were as high as 0.78 per milkweed ramet after correction for the presence of queens. Despite our observations encompassing only a limited sample across one season, the peak monarch egg densities we observed exceeded published reports from when monarch populations were higher. CONCLUSIONS: To our knowledge, this is the first study to correct for the presence of queens when calculating the density of monarch eggs. This research also provides insight into monarch utilization of less well-known regions, such as West Texas, and highlights the need to expand the scope of monarch monitoring and conservation initiatives. While the importance of monarch research and conservation in the Midwest is unquestionable, more comprehensive efforts may identify new priorities in monarch conservation and lead to a more robust and effective overall strategy, particularly given the dynamic and rapidly changing global environment.

Quantitative analysis of Northern bobwhite (Colinus virginianus) cytokines and TLR expression to eyeworm (Oxyspirura petrowi) and caecal worm (Aulonocephalus pennula) glycoproteins.

Helminth parasites have been a popular research topic due to their global prevalence and adverse effects on livestock and game species. The Northern bobwhite (Colinus virginianus), a popular game bird in the USA, is one species subject to helminth infection and has been experiencing a decline of > 4% annually over recent decades. In the Rolling Plains Ecoregion of Texas, the eyeworm (Oxyspirura petrowi) and caecal worm (Aulonocephalus pennula) helminths are found to be highly prevalent in bobwhite. While there have been increasing studies on the prevalence, pathology, and phylogeny of the eyeworm and caecal worm, there is still a need to investigate the bobwhite immune response to infection. This study utilizes previously sequenced bobwhite cytokines and toll-like receptors to develop and optimize qPCR primers and measure gene expression in bobwhite intramuscularly challenged with eyeworm and caecal worm glycoproteins. For the challenge experiments, separate treatments of eyeworm and caecal worm glycoproteins were administered to bobwhite on day 1 and day 21. Measurements of primary and secondary immune responses were taken at day 7 and day 28, respectively. Using the successfully optimized qPCR primers for TLR7, IL1ß, IL6, IFNα, IFNγ, IL10, and ß-actin, the gene expression analysis from the challenge experiments revealed that there was a measurable immune reaction in bobwhite in response to the intramuscular challenge of eyeworm and caecal worm glycoproteins.

Molecular identification of Physaloptera sp. from wild northern bobwhite (Colinus virginianus) in the Rolling Plains ecoregion of Texas.

Physaloptera spp. are common nematodes found in the stomach and muscles of mammals, reptiles, amphibians, and birds. Physaloptera spp. have a complicated life cycle with multiple definitive hosts, arthropod intermediate hosts, aberrant infections, and possible second intermediate hosts or paratenic hosts. For example, Physaloptera sp. larvae have been found within the tissues of wild northern bobwhite quail (Colinus virginianus), and it is suspected that quail may serve as paratenic or secondary hosts of these parasites. However, because it is not known what role quail play in the life cycle of Physaloptera spp. and descriptions of Physaloptera spp. larvae are limited, molecular tools may be beneficial when identifying these helminths. In this study, we generated primers using universal nematode primers and obtained a partial mitochondrial cytochrome oxidase 1 (COX 1) sequence. Morphological identification of Physaloptera sp. in bobwhite was confirmed via polymerase chain reaction (PCR), and a phylogenetic tree was constructed using the maximum likelihood method. BLAST analysis revealed a strong identity to other Physaloptera spp. and the phylogenetic tree placed all Physaloptera spp. in the same cluster. We also documented a marked increase in Physaloptera infections in bobwhite from 2017 to 2018, and the similarity of these parasites to Onchocerca volvulus and Wuchereria bancrofti may give insight into the increased prevalence we observed. This study demonstrates the usefulness of molecular techniques to confirm the identity of species that may lack adequate descriptions and provides new insight for the diagnosis and potentially overlooked significance of Physaloptera sp. infections of bobwhite in the Rolling Plains ecoregion of Texas.

Elevated parasite burdens as a potential mechanism affecting northern bobwhite (Colinus virginianus) population dynamics in the Rolling Plains of West Texas.

Northern bobwhite quail (Colinus virginianus) are a highly sought-after game bird in the Rolling Plains of West Texas. Unfortunately, bobwhite populations in this area are subject to dramatic fluctuations and have been steadily decreasing over the past several decades. While many factors have been investigated as potential mechanisms of cyclic and declining bobwhite numbers, the effect of parasites on bobwhite populations has historically been undervalued. Between December 2017 and February 2018, we received 21 hunter-shot bobwhite from Garza and Mitchell counties in Texas and found peak caecal worm (Aulonocephalus pennula) and eyeworm (Oxyspirura petrowi) burdens averaging 599 and 44, respectively. These represent the highest average parasite loads we have documented in bobwhite from the Rolling Plains thus far and are coincident with widespread reports of declining bobwhite abundance. These elevated infections also followed a high point in bobwhite populations in the Rolling Plains, and our observations of infection dynamics during this time reflect other instances of potential parasite-induced host mortality. While the sample discussed in this communication is small, our findings highlight the need for additional research into how parasites may affect bobwhite population fluctuations in this region.

Aflatoxin contamination in corn sold for wildlife feed in texas.

Supplemental feeding with corn to attract and manage deer is a common practice throughout Texas. Other species, including northern bobwhites (Colinus virginianus), are commonly seen feeding around supplemental deer feeders. In many cases, supplemental feeding continues year-round so feed supply stores always have supplemental corn in stock. Fluctuating weather and improper storage of corn can lead to and/or amplify aflatoxin contamination. Due to the recent decline of bobwhites throughout the Rolling Plains ecoregion of Texas, there has been interest in finding factors such as toxins that could be linked to their decline. In this study, we purchased and sampled supplemental corn from 19 locations throughout this ecoregion to determine if aflatoxin contamination was present in individual bags prior to being dispersed to wildlife. Of the 57 bags sampled, 33 bags (approximately 58%) contained aflatoxin with a bag range between 0.0-19.91 parts per billion (ppb). Additionally, three metal and three polypropylene supplemental feeders were each filled with 45.4 kg of triple cleaned corn and placed in an open field to study long-term aflatoxin buildup. Feeders were sampled every 3 months from November 2013-November 2014. Average concentration of aflatoxin over the year was 4.08 ± 2.53 ppb (±SE) in metal feeders, and 1.43 ± 0.89 ppb (±SE) in polypropylene feeders. The concentration of aflatoxins is not affected by the type of feeder (metal vs polypropylene), the season corn was sampled, and the location in the feeder (top, middle, bottom) where corn is sampled. It is unlikely that corn used in supplemental feeders is contributing to the bobwhite decline due to the low levels of aflatoxin found in purchased corn and long-term storage of corn used in supplemental feeders.

Human and ecological risk assessment of a crop protection chemical: a case study with the azole fungicide epoxiconazole.

Conventional risk assessments for crop protection chemicals compare the potential for causing toxicity (hazard identification) to anticipated exposure. New regulatory approaches have been proposed that would exclude exposure assessment and just focus on hazard identification based on endocrine disruption. This review comprises a critical analysis of hazard, focusing on the relative sensitivity of endocrine and non-endocrine endpoints, using a class of crop protection chemicals, the azole fungicides. These were selected because they are widely used on important crops (e.g. grains) and thereby can contact target and non-target plants and enter the food chain of humans and wildlife. Inhibition of lanosterol 14α-demethylase (CYP51) mediates the antifungal effect. Inhibition of other CYPs, such as aromatase (CYP19), can lead to numerous toxicological effects, which are also evident from high dose human exposures to therapeutic azoles. Because of its widespread use and substantial database, epoxiconazole was selected as a representative azole fungicide. Our critical analysis concluded that anticipated human exposure to epoxiconazole would yield a margin of safety of at least three orders of magnitude for reproductive effects observed in laboratory rodent studies that are postulated to be endocrine-driven (i.e. fetal resorptions). The most sensitive ecological species is the aquatic plant Lemna (duckweed), for which the margin of safety is less protective than for human health. For humans and wildlife, endocrine disruption is not the most sensitive endpoint. It is concluded that conventional risk assessment, considering anticipated exposure levels, will be protective of both human and ecological health. Although the toxic mechanisms of other azole compounds may be similar, large differences in potency will require a case-by-case risk assessment.

Validation of an enzyme immunoassay for the detection of corticosterone metabolites from northern bobwhite (Colinus virginianus) feces.

Interest in the effects of stressors on wildlife has grown substantially over the past few decades. As this interest has grown, so has the need for minimally invasive and reliable methods for estimating differences in the levels of stress hormones. An enzyme immunoassay using standardized methods was validated for detecting concentrations of corticosterone (cort) metabolites from northern bobwhite fecal samples. Two physiological challenges and one biological challenge were applied to 18 northern bobwhites (nine males and nine females), and the fecal cort metabolite concentrations were compared to baseline levels. The interactions of sex and treatment, treatment and time and sex and time were all significant. Thus, the methods and tools used here were sensitive enough to detect expected changes to the hypothalamo-pituitary-adrenal axis of northern bobwhite.

IMPLICATIONS OF TREATING PARASITIC INFECTION IN NORTHERN BOBWHITE (COLINUS VIRGINIANUS) ON OVERALL HELMINTH LIFE CYCLE.

This study was conducted to determine how reducing the parasite burden in a definitive host may affect the prevalence in intermediate hosts. Here we used the eyeworm Oxyspirura petrowi and cecal worm Aulonocephalus pennula as model species. Northern bobwhite quail (Colinus virginianus) were provided an anthelmintic medicated feed in wild systems because of convincing evidence that these parasites were suppressing their populations. Eyeworm and cecal worm prevalence were measured in Orthopterans, which act as intermediate hosts, using polymerase chain reaction. Individuals were collected from a control site, a site treated for 2 yr, and a site treated for 5 yr. Orthopteran community composition was significantly different among the sites; however, an interaction between subfamily and site was not significant for the eyeworm. There was a significant reduction in eyeworm-infected Orthopterans on the 5-yr site compared with the other 2, suggesting that treatment of a definitive host may indeed affect the prevalence of eyeworms during other life-cycle stages. There was an interaction between the Orthopteran subfamily and the site for the cecal worm, so results were analyzed within each subfamily. A significant reduction in the prevalence of cecal worms was only found in the Cyrtacanthacridinae subfamily on the 5-yr site when compared with the other sites. However, the greatest prevalence in the Gomphocerinae and Oedipodinae subfamilies across all 3 sites was 4.1%. This indicates an unknown degree of cecal worm host specificity. Therefore, conclusions could not be made through the simple assessment of prevalence.

In Vitro Lethality of Fenbendazole to the Eyeworm Oxyspirura petrowi.

Oxyspirura petrowi is a heteroxenous nematode that infects the harderian gland and other ocular tissues in birds. High-intensity infections often cause damage to the infected tissues. Due to the nature of the infection sites, treatment of O. petrowi in these hosts can be difficult. Fenbendazole (FBZ) is a common anthelmintic used to treat birds for helminth infections; however, little information exists as to the efficacy of the drug on O. petrowi infections. The present study aims to estimate lethal concentrations of FBZ to O. petrowi. Adult O. petrowi were maintained in vitro and exposed to doses of 5, 50, 100, and 200 µM concentrations of FBZ and included both negative and vehicle controls. Exposure lasted 7.5 days and lethality was determined for each treatment. Negative and vehicle controls did not differ, and both had 75% survival at the end of the treatment period. The percentage survivorship in ascending order of concentration, corrected for the controls, was 66.67%, 44.44%, 33.33%, and 0%. LC10, LC50, and LC90 estimates were 7.5 ± 0.26, 49.1 ± 1.69, and 163.2 ± 5.63 µM, respectively. In the context of known pharmacokinetics of FBZ in birds, a single oral dose of FBZ can achieve exposure levels that are lethal to O. petrowi, but the drug does not stay in the system long enough. Thus, treatment of O. petrowi infections will require multiple oral doses over several days.

MOLECULAR IDENTIFICATION OF INSECT INTERMEDIATE HOSTS WITH POTENTIAL PARASITE TRANSMISSION TO THE DEFINITIVE HOST NORTHERN BOBWHITE (COLINUS VIRGINIANUS).

Grassland birds have been declining substantially for the past several years. Habitat loss, degradation, and fragmentation as well as climate change are all thought to be the main drivers of the decline. However, as the declines continue to accelerate, it is becoming imperative to examine other factors that may contribute to population fluctuations. The nematodes Oxyspirura petrowi, Aulonocephalus pennula, and Physaloptera sp. are commonly found infecting northern bobwhite (Colinus virginianus), a game species of economic importance, and all 3 nematodes use insects as an intermediate host. Here we used polymerase chain reaction techniques to determine the occurrence of the 3 nematodes in 7 insect orders to uncover epidemiological patterns of the greatest potential for transmission to northern bobwhite. Insects were collected from March through September using sweep nets and pitfall traps. An R × C chi-squared test with Monte Carlo simulation was used to determine differences in the occurrence of the parasites across taxa and time. The results of the statistical analysis showed the nematodes are predominantly found in the order Orthoptera, and A. pennula and Physaloptera sp. showed epidemiological patterns in insects. However, no such pattern was observed with O. petrowi. An explanation for the lack of epidemiological pattern in O. petrowi is proposed and the diversity of known insect hosts of the 3 nematodes is increased.

Effects of chronic 2,4,6,-trinitrotoluene, 2,4-dinitrotoluene, and 2,6-dinitrotoluene exposure on developing bullfrog (Rana catesbeiana) tadpoles.

Chronic aqueous exposures were conducted using bullfrog (Rana catesbeiana) tadpoles (8 d old) exposed to TNT (0-4 mg/L), 2,4-DNT (0-4 mg/L), and 2,6-DNT (0-8 mg/L) for 90 d. Survival of tadpoles examined using Cox proportional hazard models was reduced at all concentrations tested. Percent of abnormal swimming and other morphological abnormalities after sublethal exposure to TNT, 2,4-DNT, and 2,6-DNT at 2 mg/L were also evaluated. The effects of TNT, 2,4-DNT, and 2,6-DNT on wet body mass, snout vent length (SVL), and developmental stage of surviving tadpoles were examined. Only 2,4-DNT did not have a significant effect on body mass or SVL, but all three compounds tested had significant effects on survival. Long-term continuous exposure to these compounds at concentrations of 0.25 mg/L could lead to significant changes in growth and survival of larval amphibians.

A Helminth Survey of Northern Bobwhite Quail (Colinus virginianus) and Passerines in the Rolling Plains Ecoregion of Texas.

The Northern bobwhite quail (Colinus virginianus) is a popular game bird that has been experiencing a well-documented decline throughout Texas since the 1960s. While much of this decline has been attributed to habitat loss and fragmentation, recent studies have identified other factors that may also contribute to decreasing quail populations. Parasites, in particular, have become increasingly recognized as possible stressors of quail, and some species, particularly the eyeworm (Oxyspirura petrowi) and cecal worm (Aulonocephalus pennula) are highly prevalent in Texas quails. Eyeworm infection has also been documented in some passerines, suggesting helminth infection may be shared between bird species. However, the lack of comprehensive helminth surveys has rendered the extent of shared infection between quail and passerines in the ecoregion unclear. Thus, helminth surveys were conducted on bobwhite, scaled quail (Callipepla squamata), Northern mockingbirds (Mimus polyglottos), curve-billed thrashers (Toxistoma curvirostre), and Northern cardinals (Cardinalis cardinalis) to contribute data to existing parasitological gaps for birds in the Rolling Plains ecoregion of Texas. Birds were trapped across 3 counties in the Texas Rolling Plains from March to October 2019. Necropsies were conducted on 54 individuals (36 quail and 18 passerines), and extracted helminths were microscopically identified. Nematode, cestode, and acanthocephalan helminths representing at least 10 helminth species were found. Specifically, A. pennula and O. petrowi had the highest prevalence, and O. petrowi was documented in all of the study species. This research adds to the body of knowledge regarding parasitic infections in quail and passerines of the Rolling Plains ecoregion and highlights the potential consequences of shared infection of eyeworms among these bird species.

Spatial distribution of lead concentrations in urban surface soils of New Orleans, Louisiana USA.

Immediately following hurricane Katrina concern was raised over the environmental impact of floodwaters on the city of New Orleans, especially in regard to human health. Several studies were conducted to determine the actual contaminant distribution throughout the city and surrounding wetlands by analyzing soil, sediment, and water for a variety of contaminants including organics, inorganics, and biologics. Preliminary investigations by The Institute of Environmental and Human Health at Texas Tech University concluded that soils and sediments contained pesticides, semi-volatiles, and metals, specifically arsenic, iron, and lead, at concentrations that could pose a significant risk to human health. Additional studies on New Orleans floodwaters revealed similar constituents as well as compounds commonly found in gasoline. More recently, it has been revealed that lead (Pb), arsenic, and vanadium are found intermittently throughout the city at concentrations greater than the human health soil screening levels (HHSSLs) of 400, 22 (non-cancer endpoint) and 390 µg/g, respectively. Of these, Pb appears to present the greatest exposure hazard to humans as a result of its extensive distribution in city soils. In this study, we spatially evaluated Pb concentrations across greater New Orleans surface soils. We established 128 sampling sites throughout New Orleans at approximately half-mile intervals. A soil sample was collected at each site and analyzed for Pb by ICP-AES. Soils from 19 (15%) of the sites had Pb concentrations exceeding the HHSSL threshold of 400 µg/g. It was determined that the highest concentrations of Pb were found in the south and west portions of the city. Pb concentrations found throughout New Orleans in this study were then incorporated into a geographic information system to create a spatial distribution model that can be further used to predict Pb exposure to humans in the city.

Molecular Identification of Oxyspirura Petrowi Intermediate Hosts by Nested PCR Using Internal Transcribed Spacer 1 (ITS1).

Recently, the heteroxenous eyeworm, Oxyspirura petrowi, has gained attention due to its prevalence in the declining game bird, Northern bobwhite (Colinus virginianus), but the intermediate hosts of many nematodes remain unknown. However, identifying the intermediate host of O. petrowi with traditional techniques would be difficult and time-consuming, especially considering there are more than 80 potential orthopteran hosts just in Texas. To screen a large number of samples quickly and effectively, primers for nested PCR (nPCR) were developed using the internal transcribed spacer 1 (ITS1) region. Then the nPCR was used to identify which of the 35 species collected from the Order Orthoptera were potential intermediate hosts of O. petrowi. With this technique, 18 potential intermediate hosts were identified. Later, we collected live specimens of species that tested positive to confirm the presence of larvae, but larvae were not found in the live specimens, nor in the extra tissue of the species that had tested positive for O. petrowi DNA. Despite this, this study demonstrated that nPCR is more sensitive than traditional techniques and can be a valuable tool in determining the intermediate hosts of parasites.

Avian and Emerging Human Oxyspirura Species Compared by Morphology, Pathogenicity, Intermediate Host, and Sequence Homology.

Based on sequence homology and phylogenetic tree results, the first report of eyeworm Oxyspirura species larvae has been confirmed in a human patient from Vietnam. However, important information related to Oxyspirura larvae was not presented in the case study. This comment provides a more detailed comparison of the Oxyspirura larvae found in the human case study to the avian eyeworm Oxyspirura petrowi.

"Weight of evidence" as a tool for evaluating disease in wildlife: An example assessing parasitic infection in Northern bobwhite (Colinus virginianus).

The potential of parasites to affect host abundance has been a topic of heated contention within the scientific community for some time, with many maintaining that issues such as habitat loss are more important in regulating wildlife populations than diseases. This is in part due to the difficulty in detecting and quantifying the consequences of disease, such as parasitic infection, within wild systems. An example of this is found in the Northern bobwhite quail (Colinus virginanus), an iconic game bird that is one of the most extensively studied vertebrates on the planet. Yet, despite countless volumes dedicated to the study and management of this bird, bobwhite continue to disappear from fields, forest margins, and grasslands across the United States in what some have referred to as "our greatest wildlife tragedy". Here, we will discuss the history of disease and wildlife conservation, some of the challenges wildlife disease studies face in the ever-changing world, and how a "weight of evidence" approach has been invaluable to evaluating the impact of parasites on bobwhite in the Rolling Plains of Texas. Through this, we highlight the potential of using "weight of the evidence" to better understand the complex effects of diseases on wildlife and urge a greater consideration of the importance of disease in wildlife conservation.

Functional PVA/VB2/TiO2 Nanofiber Webs for Controlled Drug Delivery.

Drug (vitamin B2, riboflavin, VB2)/nanoparticle (TiO2, coated on drug surface) loaded poly(vinyl alcohol) (PVA) nanofiber webs were successfully produced by the electrospinning process. The characteristics of nanofiber structure, chemical composition, mechanical properties, and drug-release properties were investigated. The morphology and diameter of nanofibers were analyzed by atomic force microscopy and scanning electron microscopy. The chemical composition and mechanical properties of nanofiber webs were examined by Fourier transform infrared spectroscopy and Instron tensile tester. The drug-release properties of nanofiber webs were studied by liquid chromatography-mass spectrometry (LC-MS/MS). The influences of TiO2:VB2 ratios (0, 18:1, 9:1, 4.5:1, 2.25:1, and 1:1) and releasing time on release behavior of PVA/VB2/TiO2 nanofiber webs were also investigated, with the corresponding virgin PVA nanofibers and LC-MS grade water as control. Among all of the tested samples, PVA/VB2/TiO2 nanofiber webs with TiO2:VB2 ratios of 18:1 and 9:1 exhibited a steady release rate with 60% of VB2 released around 168 h and all VB2 released around 10 days. In addition, by coating VB2 with TiO2, the burst release was effectively prevented due to three mechanisms: surface modification, polymer morphology, and coated surface. The results in this study indicate drug-loaded and nanoparticle-coated hydrophilic nanofiber webs are useful candidates for steady drug release in the drug delivery application field.