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.

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.

Antimicrobial Stewardship Teams in Veterans Affairs and Nonfederal Hospitals in the United States: A National Survey of Antimicrobial Stewardship Practices.

In a cross-sectional survey of US acute care hospitals, antimicrobial stewardship programs were present in most Veterans Affairs and nonfederal hospitals but varied in team composition, scope, and impact. Diagnostic stewardship was common across hospitals. Veterans Affairs hospitals had increased reach in outpatient settings. Telestewardship remains an opportunity in all hospital systems.

Transition to Oral Antibiotic Therapy for Hospitalized Adults With Gram-Negative Bloodstream Infections.

Importance: Management of gram-negative bloodstream infections (GN-BSIs) with oral antibiotics is highly variable. Objective: To examine the transition from intravenous (IV) to oral antibiotics, including selection, timing, and associated clinical and microbial characteristics, among hospitalized patients with GN-BSIs. Design, Setting, and Participants: A retrospective cohort study was conducted of 4581 hospitalized adults with GN-BSIs at 24 US hospitals between January 1 and December 31, 2019. Patients were excluded if they died within 72 hours. Patients were excluded from the oral therapy group if transition occurred after day 7. Statistical analysis was conducted from July 2022 to October 2023. Exposures: Administration of antibiotics for GN-BSIs. Main Outcomes and Measures: Baseline characteristics and clinical parameters reflecting severity of illness were evaluated in groups receiving oral and IV therapy. The prevalence of transition from IV to oral antibiotics by day 7, median day of transition, sources of infection, and oral antibiotic selection were assessed. Results: Of a total of 4581 episodes with GN-BSIs (median age, 67 years [IQR, 55-77 years]; 2389 men [52.2%]), 1969 patients (43.0%) receiving IV antibiotics were transitioned to oral antibiotics by day 7. Patients maintained on IV therapy were more likely than those transitioned to oral therapy to be immunosuppressed (833 of 2612 [31.9%] vs 485 of 1969 [24.6%]; P < .001), require intensive care unit admission (1033 of 2612 [39.5%] vs 334 of 1969 [17.0%]; P < .001), have fever or hypotension as of day 5 (423 of 2612 [16.2%] vs 49 of 1969 [2.5%]; P < .001), require kidney replacement therapy (280 of 2612 [10.7%] vs 63 of 1969 [3.2%]; P < .001), and less likely to have source control within 7 days (1852 of 2612 [70.9%] vs 1577 of 1969 [80.1%]; P < .001). Transitioning patients from IV to oral therapy by day 7 was highly variable across hospitals, ranging from 25.8% (66 of 256) to 65.9% (27 of 41). A total of 4109 patients (89.7%) achieved clinical stability within 5 days. For the 3429 episodes (74.9%) with successful source control by day 7, the median day of source control was day 2 (IQR, 1-3 days) for the oral group and day 2 (IQR, 1-4 days) for the IV group (P < .001). Common infection sources among patients administered oral therapy were the urinary tract (1277 of 1969 [64.9%]), hepatobiliary (239 of 1969 [12.1%]), and intra-abdominal (194 of 1969 [9.9%]). The median day of oral transition was 5 (IQR, 4-6 days). Total duration of antibiotic treatment was significantly shorter among the oral group than the IV group (median, 11 days [IQR, 9-14 days] vs median, 13 days [IQR, 8-16 days]; P < .001]. Fluoroquinolones (62.2% [1224 of 1969]), followed by ß-lactams (28.3% [558 of 1969]) and trimethoprim-sulfamethoxazole (11.5% [227 of 1969]), were the most commonly prescribed oral antibiotics. Conclusions and Relevance: In this cohort study of 4581 episodes of GN-BSIs, transition to oral antibiotic therapy by day 7 occurred in fewer than half of episodes, principally with fluoroquinolones, although this practice varied significantly between hospitals. There may have been additional opportunities for earlier and more frequent oral antibiotic transitions because most patients demonstrated clinical stability by day 5.

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.

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.

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.

Respiratory culture nudge improves antibiotic prescribing for Moraxella catarrhalis and Haemophilus influenzae lower respiratory tract infections.

We compared optimal antibiotic prescribing before and after implementing an interpretive ß-lactamase microbiology comment for Haemophilus influenzae and Moraxella catarrhalis in lower respiratory-tract infections. The postintervention group was associated with 5-fold increased odds of optimal de-escalation (adjusted odds ratio, 5.03; 95% confidence interval, 2.57-9.87).

Defining the Optimal Duration of Therapy for Hospitalized Patients With Complicated Urinary Tract Infections and Associated Bacteremia.

BACKGROUND: Limited data are available to guide effective antibiotic durations for hospitalized patients with complicated urinary tract infections (cUTIs). METHODS: We conducted an observational study of patients ≥18 years at 24 US hospitals to identify the optimal treatment duration for patients with cUTI. To increase the likelihood patients experienced true infection, eligibility was limited to those with associated bacteremia. Propensity scores were generated for an inverse probability of treatment weighted analysis. The primary outcome was recurrent infection with the same species ≤30 days of completing therapy. RESULTS: 1099 patients met eligibility criteria and received 7 (n = 265), 10 (n = 382), or 14 (n = 452) days of therapy. There was no difference in the odds of recurrent infection for patients receiving 10 days and those receiving 14 days of therapy (aOR: .99; 95% CI: .52-1.87). Increased odds of recurrence was observed in patients receiving 7 days versus 14 days of treatment (aOR: 2.54; 95% CI: 1.40-4.60). When limiting the 7-day versus 14-day analysis to the 627 patients who remained on intravenous beta-lactam therapy or were transitioned to highly bioavailable oral agents, differences in outcomes no longer persisted (aOR: .76; 95% CI: .38-1.52). Of 76 patients with recurrent infections, 2 (11%), 2 (10%), and 10 (36%) in the 7-, 10-, and 14-day groups, respectively, had drug-resistant infections (P = .10). CONCLUSIONS: Seven days of antibiotics appears effective for hospitalized patients with cUTI when antibiotics with comparable intravenous and oral bioavailability are administered; 10 days may be needed for all other patients.

Pharmacist-Driven Transitions of Care Practice Model for Prescribing Oral Antimicrobials at Hospital Discharge.

Importance: Although prescribers face numerous patient-centered challenges during transitions of care (TOC) at hospital discharge, prolonged duration of antimicrobial therapy for common infections remains problematic, and resources are needed for antimicrobial stewardship throughout this period. Objective: To evaluate a pharmacist-driven intervention designed to improve selection and duration of oral antimicrobial therapy prescribed at hospital discharge for common infections. Design, Setting, and Participants: This quality improvement study used a nonrandomized stepped-wedge design with 3 study phases from September 1, 2018, to August 31, 2019. Seventeen distinct medicine, surgery, and specialty units from a health system in Southeast Michigan participated, including 1 academic tertiary hospital and 4 community hospitals. Hospitalized adults who had urinary, respiratory, skin and/or soft tissue, and intra-abdominal infections and were prescribed antimicrobials at discharge were included in the analysis. Data were analyzed from February 18, 2020, to February 28, 2022. Interventions: Clinical pharmacists engaged in a new standard of care for antimicrobial stewardship practices during TOC by identifying patients to be discharged with a prescription for oral antimicrobials and collaborating with primary teams to prescribe optimal therapy. Academic and community hospitals used both antimicrobial stewardship and clinical pharmacists in a multidisciplinary rounding model to discuss, document, and facilitate order entry of the antimicrobial prescription at discharge. Main Outcomes and Measures: The primary end point was frequency of optimized antimicrobial prescription at discharge. Health system guidelines developed from national guidelines and best practices for short-course therapies were used to evaluate optimal therapy. Results: A total of 800 patients prescribed oral antimicrobials at hospital discharge were included in the analysis (441 women [55.1%]; mean [SD] age, 66.8 [17.3] years): 400 in the preintervention period and 400 in the postintervention period. The most common diagnoses were pneumonia (264 [33.0%]), upper respiratory tract infection and/or acute exacerbation of chronic obstructive pulmonary disease (214 [26.8%]), and urinary tract infection (203 [25.4%]). Patients in the postintervention group were more likely to have an optimal antimicrobial prescription (time-adjusted generalized estimating equation odds ratio, 5.63 [95% CI, 3.69-8.60]). The absolute increase in optimal prescribing in the postintervention group was consistent in both academic (37.4% [95% CI, 27.5%-46.7%]) and community (43.2% [95% CI, 32.4%-52.8%]) TOC models. There were no differences in clinical resolution or mortality. Fewer severe antimicrobial-related adverse effects (time-adjusted generalized estimating equation odds ratio, 0.40 [95% CI, 0.18-0.88]) were identified in the postintervention (13 [3.2%]) compared with the preintervention (36 [9.0%]) groups. Conclusions and Relevance: The findings of this quality improvement study suggest that targeted antimicrobial stewardship interventions during TOC were associated with increased optimal, guideline-concordant antimicrobial prescriptions at discharge.

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.

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.

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.

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.

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.

Life-cycle of Oxyspirura petrowi (Spirurida: Thelaziidae), an eyeworm of the northern bobwhite quail (Colinus virginianus).

BACKGROUND: Oxyspirura petrowi (Spirurida: Thelaziidae), a heteroxenous nematode of birds across the USA, may play a role in the decline of the northern bobwhite (Colinus virginianus) in the Rolling Plains Ecoregion of West Texas. Previous molecular studies suggest that crickets, grasshoppers and cockroaches serve as potential intermediate hosts of O. petrowi, although a complete study on the life-cycle of this nematode has not been conducted thus far. Consequently, this study aims to improve our understanding of the O. petrowi life-cycle by experimentally infecting house crickets (Acheta domesticus) with O. petrowi eggs, feeding infected crickets to bobwhite and assessing the life-cycle of this nematode in both the definitive and intermediate hosts. METHODS: Oxyspirura petrowi eggs were collected from gravid worms recovered from wild bobwhite and fed to house crickets. The development of O. petrowi within crickets was monitored by dissection of crickets at specified intervals. When infective larvae were found inside crickets, parasite-free pen-raised bobwhite were fed four infected crickets each. The maturation of O. petrowi in bobwhite was monitored through fecal floats and bobwhite necropsies at specified intervals. RESULTS: In this study, we were able to infect both crickets (n = 45) and bobwhite (n = 25) with O. petrowi at a rate of 96%. We successfully replicated and monitored the complete O. petrowi life-cycle in vivo, recovering embryonated O. petrowi eggs from the feces of bobwhite 51 days after consumption of infected crickets. All life-cycle stages of O. petrowi were confirmed in both the house cricket and the bobwhite using morphological and molecular techniques. CONCLUSIONS: This study provides a better understanding of the infection mechanism and life-cycle of O. petrowi by tracking the developmental progress within both the intermediate and definitive host. To our knowledge, this study is the first to fully monitor the complete life-cycle of O. petrowi and may allow for better estimates into the potential for future epizootics of O. petrowi in bobwhite. Finally, this study provides a model for experimental infection that may be used in research examining the effects of O. petrowi infection in bobwhite.

Identification of eyeworm (Oxyspirura petrowi) and caecal worm (Aulonocephalus pennula) infection levels in Northern bobwhite quail (Colinus virginianus) of the Rolling Plains, TX using a mobile research laboratory: Implications for regional surveillance.

Over the last few decades, there has been a decline in Northern bobwhite quail (Colinus virginianus) throughout their native range. While there are various factors that may be influencing this decline, it is suggested that parasites should be taken into consideration as a potential contributor in the Rolling Plains Ecoregion. High prevalence of the eyeworm (Oxyspirura petrowi) and caecal worm (Aulonocephalus pennula) in bobwhite of this region, coupled with a continuous decline, creates a need to assess infection through alternative methods for regional surveillance. Previous studies have developed a qPCR method and mobile research laboratory as an option for nonlethal procedures. However, there is still a need for standardization of these techniques. Therefore, this study builds on previous protocols to develop an application that considers factors that may influence qPCR results. In this study, cloacal swabs are collected from bobwhite in three locations throughout the Rolling Plains and scaled based on amount of feces present on the swab. This data is compared to qPCR standards as a limit of quantification for both eyeworm and caecal worm to define parasitic infection levels. Binary logistic regressions confirm that the probability of detection increases for both eyeworm (Odds Ratio: 2.3738; 95% Confidence Interval: [1.7804, 3.1649]) and caecal worm (Odds Ratio: 2.8516; 95% Confidence Interval: [2.2235, 3.6570]) as swab score increases. Infection levels for eyeworm and caecal worm are based on the generated cycle threshold value averages of qPCR standards. Based on the results of this study, this method can be applied in the mobile research laboratory to quantitatively assess regional parasitic infection in bobwhite throughout the Rolling Plains.

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.