Evaluation and establishment of reference intervals using the i-STAT1 blood chemistry analyzer in turkeys.

In veterinary medicine, point-of-care testing techniques have become popular, since they provide immediate results and only small amounts of blood are needed. The handheld i-STAT1 blood analyzer is used by poultry researchers and veterinarians; however, no studies have evaluated the accuracy of this analyzer determined reference intervals in turkey blood. The objectives of this study were to 1) investigate the effect of storage time on turkey blood analytes, 2) compare the results obtained by the i-STAT1 analyzer to those obtained by the GEM Premier 3000, a conventional laboratory analyzer, and 3) establish reference intervals for blood gases and chemistry analytes in growing turkeys using the i-Stat. For the first and second objectives, we used the CG8+ i-STAT1 cartridges to test blood from 30 healthy turkeys in triplicate and once with the conventional analyzer. To establish the reference intervals, we tested a total 330 blood samples from healthy turkeys from 6 independent flocks during a 3-yr period. Blood samples were then divided into brooder (<1 wk) and growing (1-12 wk of age). Friedman's test demonstrated significant time-dependent changes in blood gas analytes, but not for electrolytes. Bland-Altman analysis revealed that there was agreement between the i-STAT1 and the GEM Premier 300 for most of the analytes. However, Passing-Bablok regression analysis identified constant and proportional biases in the measurement of multiple analytes. Tukey's test revealed significant differences in the whole blood analytes between the means of brooding and growing birds. The data presented in the present study provide a basis for measuring and interpreting blood analytes in the brooding and growing stages of the turkey lifecycle, offering a new approach to health monitoring in growing turkeys.

When Is A Lineage A Species? A Case Study In Myxacorys gen. nov. (Synechococcales: Cyanobacteria) With The Description of Two New Species From The Americas.

Soil cyanobacteria are crucial components of biological soil crusts and carry out many functions in dryland ecosystems. Despite this importance, their taxonomy and population genetics remain poorly known. We isolated 42 strains of simple filamentous cyanobacteria previously identified as Pseudophormidium hollerbachianum from 26 desert locations in the North and South America and characterized these strains using a total evidence approach, that is, using both morphological and molecular data to arrive at taxonomic decisions. Based on a phylogenetic analysis of 16S rRNA gene sequences, we propose and characterize Myxacorys gen. nov. with two new species Myxacorys chilensis, the generitype, and M. californica. We also found distinct 16S-23S ITS sequence variability within species in our dataset. Especially interesting was the presence of two distinct lineages of M. californica obtained from locations in close spatial proximity (within a few meters to kilometers from each other) suggesting niche differentiation. The detection of such unrecognized lineage-level variability in soil cyanobacteria has important implications for biocrust restoration practices and conservation efforts.

A novel non-invasive method for evaluating electroencephalograms on laying hens.

The use of electroencephalograms (EEG) to study the avian brain relative to behavior was conducted as early as the 1960's. EEG readings, combined with visual cues, provide the ability to elucidate and correlate behaviors to neurological and physiological changes in a chicken. The use of EEG recordings in animal models require access to the brain to implant electrodes. Having the ability to observe EEG activity on sensible birds without surgical implantation could broaden the research in this area and give further insight related to the hen's state of awareness. The development, construction, and implementation of a minimally invasive EEG electrode placement method is described. After implementation, test animals were exposed to extreme environmental stressors as part of a concurrent depopulation methods study and EEG placement withstood the condition changes and corresponding animal physical activity. Sixteen white commercial laying hens had three monopolar 32-gauge needle electrodes inserted subcutaneously and secured to their head and body. Electrodes were attached to a pre-amplifier which transferred EEG signals to a laptop based recording system. Once the electrodes were in place, the hens were placed in individual treatment/observation chamber then various environmental stressors were applied. Verification that the observed brainwave activity was neural and not muscular was done using a photic stimulation validation test. Behavior observations were recorded to correlate sensible and insensible brainwave activity. The validation test and behavior observations demonstrated the method was successful in measuring EEG in sensible laying hens. The use of a non-surgical method for recording EEG will broaden research capabilities and enhance the understanding of a hen's response its environment, eliminate the need for invasive surgical procedures, and minimizes the confounding components of anesthesia, brain surgery, and recovery. With further refinements, the method could open new avenues in avian behavioral and physiological research.

Mucopolysaccharidosis IIIB (Sanfilippo syndrome B) in a commercial emu (Dromaius novaehollandiae) flock.

Clinicopathological diagnosis of mucopolysaccharidosis type IIIB (MPS IIIB; Sanfilippo syndrome B), an inherited autosomal recessive lysosomal storage disease, as a cause of losses in a commercial emu flock and screening breeders using a mutation-specific DNA test are described. Between 2012 and 2015, ∼5-10 juvenile emus from a few weeks to several months of age developed progressive neurological signs and died while others in the flock remained healthy. Necropsy of two affected siblings revealed multiple sites of haemorrhage, cytoplasmic periodic acid-Schiff and Luxol fast blue-positive inclusions in neurons, and aggregates of foamy macrophages in visceral organs. Affected emus were homozygous for the two-base deletion in the α-N-acetylglucosaminidase gene that causes MPS IIIB in emus. Mutation-specific DNA tests for MPS IIIB in emus were developed. Screening blood samples from 78 breeding emus revealed 14 (18%; 9 males, 4 females, and 1 unknown gender) carriers; an overall 0.09 mutant α-N-acetylglucosaminidase allele frequency. A "test and cull male carriers" programme, in which carrier males are culled but carrier females are retained, was proposed to avoid breeding affected emus together, ultimately eliminating the disease from future broods, and preserving the gene pool with as much breeding stock as possible. Molecular genetic diagnostic tests are simple, precise, and permit screening of all breeders for the mutant allele in any flock and can be used to eliminate MPS IIIB-related emu losses through informed breeding.

Molecular epidemiology of Enterococcus cecorum isolates recovered from enterococcal spondylitis outbreaks in the southeastern United States.

Enterococcus cecorum, a normal intestinal inhabitant, is increasingly responsible for outbreaks of arthritis and osteomyelitis in chickens worldwide. Enterococcal spondylitis (ES) is a specific manifestation of E. cecorum-associated disease in which increased flock morbidity and mortality result from chronic infection involving the free thoracic vertebra. In this study the genetic relatedness and antimicrobial resistance of isolates recovered from ES-affected flocks in the southeastern United States were determined. ES outbreaks from 2007 to 2011 were investigated in North Carolina (15 flocks, 13 farms, four integrators), South Carolina (one flock, one farm, one integrator) and Alabama (six flocks, six farms, one integrator). From these 22 epidemiologically distinct outbreaks, 326 isolates of E. cecorum were recovered. Isolates from spinal lesions and caeca of affected birds (cases) and caeca of unaffected birds (controls) were genotyped using pulsed-field gel electrophoresis; phenotyped using both GenIII MicroPlate™ (Biolog; Hayward, CA, USA) microbial identification plates and antimicrobial sensitivity testing; and compared with each other. Isolates from spinal lesions were incapable of mannitol metabolism and the majority of these isolates were genetically clonal. In contrast, caecal isolates from control birds varied in their ability to metabolize mannitol and were genetically diverse. Isolates from both case and control birds had high levels of antimicrobial resistance. These findings indicate that the increase in E. cecorum-associated disease in the southeast United States is due to the emergence of new clones with increased pathogenicity and multidrug resistance.

An outbreak and source investigation of enterococcal spondylitis in broilers caused by Enterococcus cecorum.

Enterococcus cecorum was isolated from spondylitis lesions in broilers from two flocks in North Carolina that were experiencing increased mortality. Affected birds showed paresis and paralysis, clinical signs characteristic of enterococcal spondylitis (ES). Affected birds rested on their hocks and caudal abdomens with legs extended forward and were unable to stand or walk. Necropsy examination of affected birds revealed firm to hard inflammatory masses involving the vertebral bodies at the level of the free thoracic vertebra that bulged dorsally and compressed the spinal cord. When opened, lesions contained pale, tan to yellow caseonecrotic material. Microscopically, necrosis and fibrinoheterophilic spondylitis with intralesional gram-positive bacteria were seen. Heavy growth of E. cecorum recovered from vertebral lesions confirmed the diagnosis of ES. To investigate possible sources of the organism for one of the flocks bacterial cultures were made from the environment, water lines, mice trapped on the farm, cecal/cloacal swabs from one of the parent broiler breeder flocks, egg residue, hatching eggs, and the hatchery environment. Except for cecal/cloacal swabs from the breeders, E. cecorum was not isolated from any of these samples. When compared phenotypically and genotypically, cecal/cloacal isolates of E. cecorum from the breeders differed from isolates from spondylitis lesions in the broilers. The source of E. cecorum for the broiler flocks was not determined, but vertical transmission appears unlikely.

Selected blood chemistry values in mobility-impaired broiler breeder hens with suspected calcium tetany using the i-STAT handheld clinical analyzer.

Calcium tetany is a poorly defined disease of broiler breeder hens that results from acute hypocalcemia. It is characterized by impaired mobility, increased mortality, and absence of gross lesions that would explain the impaired mobility. To evaluate if hens with impaired mobility had calcium tetany or other abnormalities, blood values from normal and affected hens were determined using the i-STAT handheld clinical analyzer. Three flocks were evaluated weekly prior to peak production (range 25-30 wk of age) comparing normal hens to hens with clinically apparent calcium tetany. Calcium tetany suspect (CaTS) hens from four additional flocks were also evaluated. Significant hypocalcemia (P < 0.001) was observed in CaTS hens (average = 1.14 mmol/L ionized calcium [iCa]) compared to normal hens (average = 1.53 mmol/L iCa) in only one of three flocks sampled weekly. Clinically affected hens from one of the other four flocks also had hypocalcemia. Blood value abnormalities in mobility-impaired hens without hypocalcemia included hypernatremia. Findings in this study indicate calcium tetany is one cause of impaired mobility in breeder hens, but mobility impairment without hypocalcemia can also occur. Calcium tetany should be confirmed by finding significantly decreased levels of iCa in the blood, as diagnosis based on clinical presentation and necropsy results can be inaccurate. The i-STAT handheld clinical analyzer is an efficient, relatively low-cost method to determine iCa and other blood chemistry values that may be associated with impaired mobility in broiler breeder hens.

Experimental reproduction of enterococcal spondylitis in male broiler breeder chickens.

There has been a recent emergence of epidemic spinal infections with necrosis causing lameness and mortality in male broilers and broiler breeders. Mortality in affected flocks may be as high as 15%. The disease has been called enterococcal spondylitis (ES), based on the frequent isolation of Enterococcus cecorum from the lesions and necrosis and inflammation observed in the free thoracic vertebrae (FTV) of affected birds. Male broiler breeders in an experimental setting were challenged with pure E. cecorum isolates obtained from ES-affected commercial flocks. Challenge routes included oral gavage (10(8)), intravenous (i.v.; 10(3)), and air sac (AS; 10(3)). Half the study birds in each group were chemically immunosuppressed with dexamethasone. Spinal lesions were observed grossly in birds challenged intravenously (2.9%) and birds challenged orally (6.1%). Microscopic spinal lesions consistent with ES were more frequently identified compared with gross lesions in the orally challenged group (30.3%). Chemical immunosuppression with dexamethasone was not associated with a greater incidence of ES in this study. By recreating the disease experimentally, the study design reported here may help in the further development of an experimental challenge model for future studies on risk factors, prevention, and therapeutic intervention of ES.

Selected blood chemistry and gas reference ranges for broiler breeders using the i-STAT handheld clinical analyzer.

Selected blood chemistry and gas reference ranges for clinically healthy broiler breeder hens were established using CG8+ cartridges in an i-STAT handheld point-of-care clinical analyzer. Samples from 165 hens (25-36 wk of age), representing three broiler breeder strains reared by four integrators, were evaluated. A standardized sampling technique was developed to minimize instrument error readings. The following reference ranges and means, respectively, were determined: sodium (141.6-152.6, 147.1 [mmol/L]), potassium (4.1-5.7, 4.9 [mmol/L]), ionized calcium (1.20-1.73, 1.47 [mmol/L]), glucose (207.2-260.7, 234.0 [mg/dl]), hematocrit (21.3-30.8, 26.1 [% packed cell volume]), hemoglobin (7.3-10.5, 8.9 [g/dl]), pH (7.28-7.57, 7.42), carbon dioxide partial pressure (25.9-49.5, 37.7 [mm Hg]), oxygen partial pressure (32.0-60.5, 46.2 [mm Hg]), bicarbonate (18.9-30.3, 24.6 [mmol/L]), total carbon dioxide (19.9-31.5, 25.7 [mmol/L]), base excess (-6.8 to 7.2, 0.2), and oxygen saturation (70.6-93.3, 82.0 [%]). Wide ranges in blood gases and base excess occurred in all strains. Cobb strain hens had significantly lower glucose and higher partial and saturated oxygen values compared with two Ross strains. Significant differences in several blood parameters were found among different integrators and in older postpeak production birds. The i-STAT handheld point-of-care clinical analyzer provides rapid, relatively low cost, blood chemistry values that are useful for investigating broiler breeder flock diseases of unknown or uncertain etiology, especially those suspected of having a metabolic cause.

Prevalence and distribution of Salmonella in organic and conventional broiler poultry farms.

The objective of this cross-sectional study was to compare the prevalence of Salmonella and antimicrobial-resistant Salmonella, as well as investigate the distribution of this pathogen in organic and conventional broiler poultry farms. Fecal (n = 420), feed (n = 140), and drinking water (n = 140) samples were collected from birds at 3 and 8 weeks of age for 2-flock cycles. One house was sampled per farm at three organic and four conventional broiler farms from the same company in North Carolina. All samples were analyzed for the presence of Salmonella using selective enrichment techniques. Further phenotypic (antimicrobial susceptibility) and genotypic (pulsed-field gel electrophoresis [PFGE]) testing were performed. Salmonella prevalences in fecal samples were 5.6% (10/180) and 38.8% (93/240) from organic and conventional farms, respectively. From feed, 5.0% (3/60) and 27.5% (22/80) of the samples were positive for Salmonella from organic and conventional farms, respectively. None of the water samples were positive for Salmonella. Seventy isolates were characterized by antimicrobial susceptibility and PFGE types. The two most common resistance phenotypes were single resistance to streptomycin (36.2% [25/58]: conventional; 25% [3/12] organic), and multidrug resistance to six antimicrobial agents: ampicillin-streptomycin-amoxicillin/clavulanic acid-cephalothin-ceftiofur-cefoxitin (AmStAxChCfFx; 39.7%: conventional only). Genotypic analysis using PFGE showed clonality among isolates within and between the two types of farms. The results of our study suggest that within this poultry company, the prevalence of fecal Salmonella was lower in certified-organic birds than in conventionally raised birds, and the prevalence of antimicrobial-resistant Salmonella was also higher in conventionally raised birds than in certified-organic birds.

Preparedness and disaster response training for veterinary students: literature review and description of the North Carolina State University Credentialed Veterinary Responder Program.

The nation's veterinary colleges lack the curricula necessary to meet veterinary demands for animal/public health and emergency preparedness. To this end, the authors report a literature review summarizing training programs within human/veterinary medicine. In addition, the authors describe new competency-based Veterinary Credential Responder training at North Carolina State University College of Veterinary Medicine (NCSU CVM). From an evaluation of 257 PubMed-derived articles relating to veterinary/medical disaster training, 14 fulfilled all inclusion requirements (nine were veterinary oriented; five came from human medical programs). Few offered ideas on the core competencies required to produce disaster-planning and response professionals. The lack of published literature in this area points to a need for more formal discussion and research on core competencies. Non-veterinary articles emphasized learning objectives, commonly listing an incident command system, the National Incident Management System, teamwork, communications, and critical event management/problem solving. These learning objectives were accomplished either through short-course formats or via their integration into a larger curriculum. Formal disaster training in veterinary medicine mostly occurs within existing public health courses. Much of the literature focuses on changing academia to meet current and future needs in public/animal health disaster-preparedness and careers. The NCSU CVM program, in collaboration with North Carolina Department of Agriculture and Consumer Service, Emergency Programs and University of North Carolina at Chapel Hill School of Public Health, operates as a stand-alone third-year two-week core-curriculum training program that combines lecture, online, experiential, and group exercises to meet entry-level federal credentialing requirements. The authors report here its content, outcomes, and future development plans.

Bacterial orchitis and epididymo-orchitis in broiler breeders.

Two cases of orchitis and epididymo-orchitis in broiler breeders are described. The first case occurred in a 62-week-old rooster showing signs of breeding inactivity. Grossly, the left testis was abnormally shaped and had multiple yellow foci, prominent blood vessels, and a gelatinous mass on the surface. The right testis and other tissues appeared normal. A pure, heavy growth of Staphylococcus aureus was obtained on bacterial culture, and intralesional Gram-positive cocci were numerous on histopathology. The second case occurred in a 28-week-old rooster that was found dead. Both testes were abnormally shaped, swollen, firm, and had irregular, dark, depressed areas, prominent vessels, and multiple petechial haemorrhages. Epididymides were enlarged and nodular. Other lesions in this rooster included poor nutritional condition, distended cloaca, and urate scalding of the abdominal skin below the vent. Microscopically, there was extensive, severe heterophilic intratubular orchitis and epididymitis with intralesional Gram-negative bacteria. A heavy, pure growth of Escherichia coli was obtained from the testes and epididymides. In both cases, the findings suggested that the most probable route of infection was ascending via the ductus deferens.

A laboratory exercise to determine human ABO blood type by noninvasive methods.

Analysis of single-nucleotide polymorphisms and their association with diseases and nondisease phenotypes is of growing importance in human biology studies. In this laboratory exercise, students determine the genetic basis for their ABO blood type; however, no blood is drawn. Students isolate genomic DNA from buccal mucosa cells that are present in saliva and analyze the DNA on an agarose gel. Subsequently, this DNA is used as a PCR template to amplify exons 6 and 7 of the gene that determines the human ABO phenotype. These PCR products are digested and run on agarose gels to examine the restriction fragment length polymorphisms. A deletion in the O(1) allele converts the BstE II site in exon 6 into a Kpn I site, and this feature is used to determine the presence of O(1) alleles. The pattern of exon 7 digest products allows students to distinguish among four other common ABO alleles: A(1) , A(2) , B, and O(2) . This exercise introduces students to commonly used molecular biology techniques, such as genomic DNA isolation, PCR, gel electrophoresis, and restriction digests.

Evaluation of the effectiveness of two infectious bronchitis virus vaccine programs for preventing disease caused by a California IBV field isolate.

Infectious bronchitis virus CA99 serotype was isolated from several broiler flocks in Northern California. The virus caused late-onset respiratory disease and increased airsacculitis condemnation in affected flocks despite the use of an established infectious bronchitis virus vaccination program. An experimental study compared Holland/Arkansas and Massachusetts/Arkansas vaccination protocols to determine the efficacy of commercial infectious bronchitis virus vaccines in reducing respiratory disease and airsacculitis lesions found at processing that were associated with a CA99 field isolate. All vaccination groups were given Massachusetts/Connecticut strains of infectious bronchitis virus vaccines at age 1 day followed by vaccination with either Holland/ Arkansas or Massachusetts/Arkansas vaccine strains at 18 days of age. Birds were challenged at age 31 days with a CA99 field isolate. Gross pathology, histopathology, and virus isolation were evaluated. Chickens vaccinated with Holland/Arkansas had marginally better protection against CA99 challenge than chickens vaccinated with Massachusetts/Arkansas, although differences were not statistically significant.

Disseminated Aspergillus flavus infection in broiler breeder pullets.

Increased morbidity and mortality occurred in a 5-wk-old broiler breeder replacement pullet flock. The affected broiler pullet flock was housed on the first floor of a two-story confinement building. Mortality increased to 0.1%/day compared to the flock on the second floor, which had mortality levels of less than 0.01%/day. Clinical signs in the affected chickens included inactivity, decreased response to stimuli, and anorexia. No respiratory or neurologic signs were observed. On necropsy, affected pullets were dehydrated and emaciated and had disseminated variably sized single or multiple heterophilic granulomas that contained intralesional septate and branching fungal hyphae. Lesions were extensive around the base of the heart in the thoracic inlet and in the kidneys. Other affected organs included eyelid, muscle, proventriculus, ventriculus, intestine, liver, spleen, lung, and heart. Aspergillus flavus was cultured from the visceral granulomas. The source of flock exposure to the organism was not determined.