Evaluation of mechanical cervical dislocation, captive bolt, carbon dioxide, and electrical methods for individual on-farm euthanasia of broiler breeders.

Efficacious euthanasia by applying manual cervical dislocation can be difficult on large and mature poultry. The challenge with using manual cervical dislocation is that the strength required to hold heavy poultry and swiftly apply cervical dislocation can be physically impossible for most people. Therefore, alternative methods of euthanasia are needed for mature and large poultry. Mechanical cervical dislocation using the Koechner Euthanizing Device (KED), captive bolt using the Turkey Euthanasia Device (TED), carbon dioxide (CO2), and electrical euthanasia were evaluated for use on 65-wk-old broiler breeders at flock termination. Following application of each method, physiological reflexes including the eye nictitating membrane reflex, mouth gaping, and body movement, broken skin, blood loss, kill success, time to cessation of heartbeat, and blood plasma corticosterone levels were assessed. Birds euthanized using the KED had longer response durations for eye nictitating membrane (91 s) and reflexive mouth gaping (161 s) compared to TED, CO2, and electrical euthanasia (0-7 s). Body movement durations were also longer for KED (214 s) and TED (209 s) than for CO2 and electrical euthanasia (0-8 s). The highest percentages of broken skin (93%) and blood loss (96%) were observed for TED, followed by KED (71%, 68%), then CO2 (0%, 6%) and electrical euthanasia (0%, 3%). No significant differences (P = 0.1781) were observed for kill success rates with 98% for KED, 100% for TED, 97% for CO2, and 100% for electrical euthanasia at 4-min. Time to heartbeat cessation did not differ between KED (659 s), TED (427 s), or CO2 (583 s) euthanasia methods. No heartbeat was detected following electrical euthanasia. Blood plasma corticosterone levels did not differ between preeuthanasia or posteuthanasia from any of the methods applied. Based on these results each euthanasia method is acceptable for use with broiler breeders.

Assessment of Stabilized Hydrogen Peroxide for Use in Reducing Campylobacter Levels and Prevalence on Broiler Chicken Wings.

ABSTRACT: Poultry processing establishments use antimicrobial aids on broiler parts to minimize Campylobacter contamination. A silver-stabilized hydrogen peroxide (SHP) product was assessed for use as an antimicrobial processing aid. In a series of experiments, wing segments with skin were inoculated with 103 to 107 cells of Campylobacter coli, followed by treatment with SHP at 15,000 or 30,000 mg/L, peroxyacetic acid (PAA) at 300 or 3,000 mg/L (parts per million), or water. Each treatment was applied by either dip or spray. Rinsates from each wing segment were analyzed for direct counts and prevalence of Campylobacter. Treatment with SHP or PAA significantly reduced Campylobacter levels compared with water controls by up to 2.22 log CFU/mL. At high inoculum levels (106 to 107), SHP and PAA applied by dip had up to 1.27 log CFU/mL further reductions of Campylobacter levels compared with spray-treated wing segments. Additionally, wing drumettes were observed to retain higher levels and prevalence of Campylobacter recovery compared with wing flats at a low inoculation level (103). The results indicated that there was no carryover effect of SHP (same day versus 24 h) and dip treatment with SHP or PAA decreased Campylobacter recovery on broiler chicken wing segments compared with a water control. Although a 2-log reduction was modest, SHP had similar efficacy as the commonly used processing aid PAA. SHP shows potential for further investigation as an antimicrobial processing aid for use on poultry parts.

Impact of alternative electrical stunning parameters on the ability of broilers to recover consciousness and meat quality.

Broilers in the United States are typically electrically stunned using low voltage-high frequency (12-38 V, ≥400 Hz) DC or AC water bath stunners. In the European Union, however, broilers are required to be electrocuted using high voltage-low frequency (50-150 V, 50-350 Hz) AC. Low voltage stunned broilers regain consciousness in the absence of bleeding. In contrast, high voltage stunned broilers die due to induction of cardiac fibrillation. For birds stunned with low voltage systems, concerns have been raised regarding animal welfare during bleeding. This work evaluated the impact of extended DC stunning duration and alternative stunning methods (DC+AC combination) on the recovery of bird consciousness and meat quality. In the absence of bleeding, broilers that were DC stunned for extended times (60, 90, or 120 s), 63, 10, or 0% of broilers, respectively, were able to recover consciousness. Alternative stunning protocols included water bath stunning broilers at 15 or 25 V DC for 10 s followed by plate stunning at 100, 110, or 120 V AC for 5 s. Prior to shackling, live body weight and shank width were measured and during stunning, maximum mA for both DC and AC stuns were recorded. All of the alternative stunning protocols (DC+AC) resulted in non-recoverable stunning. The maximum mA recorded during both DC and AC stunning were moderately/strongly (r = 0.54-0.81) correlated to body weight and poorly/moderately (r = 0.27-0.74) correlated to shank width. No significant differences for carcass or meat quality characteristics (hemorrhages, red wing tips, broken clavicles, pH, cook loss, a* and b* color values, and MORS shear energy) were detected between control (15 or 25 V DC only) and treatment groups (DC+AC combination stunning). The only significant different meat quality parameter was L* values where the lowest voltage group (15 V DC) had the darkest fillets (53.27) and the 15 V DC+100 V AC group had the lightest fillets (55.61) with all other groups intermediate. These data indicate that stunning parameters combining DC and AC stunning may be viable protocols when a stun-to-death is desired.

Carcass orientation and drip time affect potential surface water carryover for broiler carcasses subjected to a post-chill water dip or spray1.

To estimate the potential for residual antimicrobial solution carryover, surface water accumulation and loss was measured on post-chill carcasses that were either dipped or sprayed with water. For all experiments, broilers were slaughtered, soft or hard scalded, defeathered, and eviscerated. Carcasses were immersion chilled, allowed to drip, and post-chill carcass weight (CW) recorded. For water dip treatment, carcasses were dipped for 0.5 min in water and hung by a wing (n = 33) or a leg (n = 30) and CW recorded at 0, 0.5, 1, 2, and 5 min post-dip. For water spray treatment, individual carcasses were hung by either the wings (n = 35) or legs (n = 34) from a shackle suspended from a scale. Water was sprayed at 80 psi and post-spray CW recorded. Initial water accumulation (0 min) for dipped carcasses was not significantly different (P > 0.05) for carcasses hung by the leg (101.0 g) or wing (108.8 g). Following the 5 min drip time, 31 g of water remained on the carcasses hung by the leg and only 10 g on carcasses hung by the wing (P < 0.05). When carcasses were sprayed with water, initial water accumulation (0 min) was 62 g for carcasses hung by the legs and 60 g for carcasses hung by the wings (P > 0.05). Following the 5 min drip time, 1 g or no water remained on the sprayed carcasses (P > 0.05). Carcasses that were dipped and hung by a leg for 5 min retained significantly more water (31 g) than carcasses that were dipped and hung by a wing (10 g) or sprayed carcasses hung either way (0.3 g) (P < 0.05). Post-chill water dip resulted in significantly higher initial carcass water accumulation than spraying (105 g vs. 61 g, P < 0.05). Carcass orientation during dripping only affected the amount of retained water for dipped carcasses. Dipped carcasses hung by a leg have the highest potential for residual carcass antimicrobial solution carryover and sprayed carcasses hung by either orientation have the lowest potential for residual antimicrobial solution carryover.

Mesenchymal stem cells from the retropatellar fat pad and peripheral blood stimulate ACL fibroblast migration, proliferation, and collagen gene expression.

Mesenchymal stem cells (MSCs) have been of recent interest as adjuncts for ligament repair. However, the effect of these cells on the resident ligament fibroblasts has not yet been defined. In this study, we hypothesized that co-culture of MSCs and ligament fibroblasts would result in increases in the proliferative rate of the ligament fibroblasts and their expression of collagen-related genes, as well as differentiation of the MSCs down a fibroblastic pathway. In addition, we hypothesized that these effects would be dependent on the source of the MSCs. Porcine MSCs were isolated from both the retro-patellar fat pad (ADSCs) and the peripheral blood (PBMCs) and co-cultured with porcine anterior cruciate ligament (ACL) fibroblasts. Fibroblast migration, proliferation, and collagen gene expression were evaluated at time points up to 14 days. ADSCs had a greater effect on stimulating ACL-fibroblast proliferation and procollagen production, while PBMCs were more effective in stimulating ligament fibroblast migration. In addition, co-culture with the ACL fibroblasts led to significant increases in collagen gene expression for ADSCs, suggesting a differentiation of these cells down a fibroblastic pathway during the co-culture period. This was not seen for the PBMCs. Thus, the effects of MSCs on in situ ACL fibroblasts were found to be source dependent, and the choice of MSC source should take into account the different performance characteristic of each type of MSC.

Recruitment bias resulted in poorer overall health status in a community-based control group.

OBJECTIVE: During a prospective community-based incidence study of parkinsonism, a control group was recruited for comparison with the incident patients. This study compared the demographic and health status of recruited vs. nonrecruited controls. STUDY DESIGN AND SETTING: For each incident patient, attempts were made to recruit an age-gender matched control from the same general practice or, failing that, from a previously identified community cohort of people aged over 64 years who had expressed an interest in taking part in future research. Recruited controls were compared with those who were approached but not recruited in terms of age, socioeconomic status, gender, several measures of health status, and survival. RESULTS: A total of 74 controls (40%) were recruited out of 186 potential controls who were approached. Recruited controls scored slightly worse than nonrecruited controls on every measure of health status, which reached statistical significance for numbers of acute prescriptions and major surgical procedures. There were no significant differences in age, gender, socioeconomic status, or survival. CONCLUSION: The control cohort was affected by recruitment bias, which suggested that recruited controls had slightly poorer health compared to nonrecruited controls. This bias may reduce differences in health when comparisons are made between the controls and the parkinsonian patients.

Changes in diagnosis with follow-up in an incident cohort of patients with parkinsonism.

BACKGROUND: Accurate diagnosis of the cause of parkinsonism during life can be difficult, particularly at presentation, but few studies have described changes in clinical diagnosis over time and the effect of applying strict research criteria. METHODS: Incident patients with a possible/probable diagnosis of degenerative or vascular parkinsonism had a standardised assessment at diagnosis and at yearly intervals thereafter at which the most likely clinical diagnosis was recorded without strict application of research criteria. Four years after the beginning of the incident period, formal research criteria were applied retrospectively using patient records at baseline and the latest yearly follow-up. RESULTS: Of 82 incident patients, 66 underwent at least 1 year of follow-up. After a median follow-up of 29 months, clinical diagnosis had changed in 22 (33%). Most (82%) changes occurred in the first year and were due to the development of atypical clinical features, particularly early cognitive impairment; the results of brain imaging; responsiveness to levodopa; and the rate of disease progression. Diagnosis on research criteria differed from latest clinical diagnosis in eight participants (12%). Research criteria gave a "probable" diagnosis in 71% of parkinsonian patients at follow-up but in only 15% at the initial assessment. DISCUSSION: The clinical diagnosis of the cause of parkinsonism at presentation was often incorrect, even when made by those with a special interest. In particular, Parkinson's disease was overdiagnosed. Research criteria were often unhelpful in clarifying the diagnosis, even after a median of 29 months of follow-up. Further research is required to identify factors that may be used to improve the accuracy of diagnosis at initial assessment.

Changes in quality of life in people with Parkinson's disease left untreated at diagnosis.

BACKGROUND: The issue of whether to adopt a "wait and watch" strategy or to initiate drug therapy soon after diagnosis in Parkinson's disease (PD) has been the subject of some debate. A recent observational study supported early treatment by demonstrating deterioration in self-reported health status in those left untreated, but not those who received therapy. We aimed to replicate this observation. METHODS: People with PD from a prospective incidence study underwent follow-up with yearly clinical assessment of parkinsonian impairment (Unified Parkinson's Disease Rating Scale (UPDRS)) and self-reported health status (Parkinson's Disease Questionnaire (PDQ-39)). Two year outcomes were compared with those who started treatment within 1 year of diagnosis and those left untreated. RESULTS: 42 patients with PD were followed-up for 2 years, of whom 26 started treatment during the first year and 16 remained untreated. Those receiving treatment had significantly higher UPDRS and PDQ-39 scores at baseline. There was no significant deterioration in PDQ-39 score in either group (median change untreated 0.8 vs treated 4.0; p = 0.47), despite a significant difference in the change in motor UPDRS scores (untreated 6.0 vs treated -6.0; p = 0.03). CONCLUSION: Given the lack of significant deterioration in the PDQ-39 in untreated patients, we believe a "wait and watch" strategy for the treatment of newly diagnosed PD remains a credible approach unless randomised trials prove otherwise.

Screening for undiagnosed parkinsonism in people aged 65 years and over in the community.

We screened a random sample of 2449 people aged 65 years and over for undiagnosed parkinsonism, using a postal screening questionnaire followed by clinical neurological assessment. Amongst the 1556 (63.5%) patients who responded, four patients with previously undiagnosed parkinsonism were identified, suggesting a prevalence of 257 per 100,000 (95% CI 70, 658) in this age-group. Although only small, the numbers were sufficient to significantly increase the incidence of parkinsonism in an incidence study. Two simple screening questions achieved a high sensitivity for newly diagnosed parkinsonism of 95%, but a low specificity of 28%.

The use of the Internet to access oral health-related information by patients attending dental hygiene clinics.

BACKGROUND: The degree to which patients access the Internet for oral health-related information is unknown. OBJECTIVES: This study was designed to determine if patients attending student dental hygiene clinics had: (1) accessed the Internet, (2) used the Internet as a source of information on both general health and oral health, (3) bought oral health care products on-line, and (4) were interested in being directed to appropriate on-line oral health resources by dental professionals. METHODS: A self-complete questionnaire was administered to a quota sample of 280 new and returning patients attending the student dental hygiene clinic at either the Cardiff Dental Hospital or a nearby Primary Dental Care Unit. RESULTS: A total of 269 (96%) patients consented to participate in the study. Of these, 147 (54.6%) had accessed the Internet, the majority (61.2%) from home. Internet use was significantly related to both patient age and level of education (P < 0.001). In total, 59 patients claimed accessing the Internet for information on general health topics. Only eight respondents had used the Internet for information on oral health/dental topics, with seven reporting that they found the information useful. On-line purchase of oral health products was reported by seven patients. However, 117 (43.5% of all respondents) patients indicated that they would be interested in the Internet as a future source of oral health information and 112 (41.6%) would appreciate being directed to appropriate sites by their dentist or dental hygienist. CONCLUSIONS: Patients attending student dental hygiene clinics make little use of the Internet for oral health information. However, reported interest in future access is such that dental professionals should develop and harness the potential of the Internet as an educational resource.

To solve a deadly shortage: economic incentives for human organ donation.

In this article Dr. Harris and attorney Alcorn propose the establishment of a governmentally regulated, posthumous organ market, with economic incentives for the donors, in order to increase the supply of transplantable organs. The authors review transplant technology, provide a short history of donation and sale of organs, tissues, and cells, discuss the various legislative approaches that have been made to increase the supply of organs, and analyze the problems with the open market approach. They conclude with a proposal for a regulated posthumous organ market.

Biochemical and biological characterization of a human Rac2 GTPase mutant associated with phagocytic immunodeficiency.

The Rho GTPase, Rac2, is expressed only in hematopoietic cell lineages, suggesting a specific cellular function in these cells. Genetic targeting studies in mice showed that Rac2 is an essential regulator of neutrophil chemotaxis, L-selectin capture and rolling, and superoxide production. Recently, a dominant negative mutation of Rac2, D57N, has been reported to be associated with a human phagocytic immunodeficiency. To understand further the cellular phenotypes associated with this D57N Rac2 mutant we examined its biochemical characteristics and functional effects when expressed in primary murine bone marrow cells. When compared with wild type (WT) Rac2, D57N Rac2 displayed approximately 10% GTP binding ability resulting from a markedly enhanced rate of GTP dissociation and did not respond to the guanine nucleotide exchange factors. These results suggest that D57N Rac2 may act in a dominant negative fashion in cells by sequestering endogenous guanine nucleotide exchange factors. When expressed in hematopoietic cells, D57N Rac2 reduced endogenous activities of not only Rac2, but also Rac1 and decreased cell expansion in vitro in the presence of growth factors due to increased cell apoptosis. Unexpectedly, D57N expression had no effect on proliferation. In contrast, expansion of cells transduced with WT Rac2 and a dominant active mutant, Q61L, was associated with significantly increased proliferation. Transplantation of transduced bone marrow cells into lethally irradiated recipients showed that the percentage of D57N-containing peripheral blood cells decreased markedly from 40% at 1 month to <5% by 3 months postinjection. Neutrophils derived in vitro from the transduced progenitor cells containing D57N demonstrated markedly impaired migration and O(2)(-) responses to formyl-methionyl-leucyl-phenylalanine, reflecting the same cellular phenotype in these differentiated cells as those described previously in patient cells. These data suggest that the phenotypic abnormalities associated with D57N Rac2 may involve not only neutrophil cellular functions, but also abnormal cell survival in other hematopoietic cells and that overexpression of Rac leads to increased proliferation of normal cells in vitro, whereas deficiency of Rac leads to increased apoptosis.

Immunosuppressed surfactant protein A-deficient mice have increased susceptibility to Pneumocystis carinii infection.

Immunosuppressed Swiss Black mice deficient in surfactant protein A (SP-A(-/-)) and wild-type control mice (SP-A(+/+)) were exposed to Pneumocystis carinii by environmental exposure, intratracheal inoculation, and direct exposure to other infected animals. The frequency and intensity of P. carinii infection were significantly greater in the SP-A(-/-) mice by all 3 methods of exposure. P. carinii free of SP-A and alveolar macrophages were isolated from SP-A(-/-) mice and were tested in an in vitro attachment assay. Pretreatment of P. carinii with human SP-A resulted in a significant dose-dependent increase of the adherence of P. carinii to the macrophages. Thus, SP-A plays a role in host defense against P. carinii in vivo, perhaps by functioning as a nonimmune opsonin.

A new compound heterozygous mutation of the gonadotropin-releasing hormone receptor (L314X, Q106R) in a woman with complete hypogonadotropic hypogonadism: chronic estrogen administration amplifies the gonadotropin defect.

We describe a woman with complete hypogonadotropic hypogonadism and a new compound heterozygous mutation of the GnRH receptor (GnRHR) gene. A null mutation L314X leading to a partial deletion of the seventh transmembrane domain of the GnRHR is associated with a Q106R mutation previously described. L314X mutant receptor shows neither measurable binding nor inositol phosphate production when transfected in CHO-K1 cells compared to the wild-type receptor. The disease is transmitted as an autosomal recessive trait, as shown by pedigree analysis. Heterozygous patients with GnRHR mutations had normal pubertal development and fertility. The present study shows an absence of LH and FSH response to pulsatile GnRH administration (20 microg/pulse, sc, every 90 min). However, GnRH triggered free alpha-subunit (FAS) pulses of small amplitude, demonstrating partial resistance to pharmacological doses of GnRH. FSH, LH, and FAS concentrations were evaluated under chronic estrogen treatment and repeat administration of GnRH. Not only were plasma FSH, LH, and FAS concentrations decreased, but FAS responsiveness was reduced. This new case emphasizes the implication of the GnRH receptor mutations in the etiology of idiopathic hypogonadotropic hypogonadism. We also have evidence for a direct negative estrogen effect on gonadotropin secretion at the pituitary level, dependent on the GnRHR signaling pathway.

Development of photosystem-II activity during irradiance of etiolated Helianthus (Asteraceae) seedlings.

Light-induced development of photosystem (PS)-II activity was followed during irradiance of etiolated Helianthus annuus (sunflower) cotyledons using chlorophyll a fluorescence. Cotyledons from seedlings grown in continuous darkness for 6 d were exposed to 100 µmol photons·m(-2)·s(-1) for time periods of 1, 3, 6, and 12 h. Associated with increased time of irradiance exposure were significant: (1) increases in concentration of PS II, (2) increases in quantum efficiency of PS II, (3) decreases in the ratio of PS-II quinone(B) (Q(B))-nonreducing centers to total PS-II centers (PS-II Q(B)-nonreducing centers + PS-II Q(B)-reducing centers), and (4) decreases in the ratio of slow PS-II Q(B)-reducing centers to total PS-II Q(B)-reducing centers (fast PS-II Q(B)-reducing centers + slow PS-II Q(B)-reducing centers). The results support the hypotheses that development of PS II involves assembly of complexes which initially cannot reduce Q(B) and that heterogeneous aspects of PS-II pools during chloroplast maturation may represent different developmental states.

A novel heterogeneous nuclear ribonucleoprotein-like protein interacts with NS1 of the minute virus of mice.

NS1, the major nonstructural parvovirus protein of the minute virus of mice, is a multifunctional protein responsible for several aspects of viral replication. NS1 transactivates the P38 promoter (used to express the structural proteins), as well as its own strong promoter, P4. To study the mechanism of activation and to map regions of NS1 responsible for transactivation, NS1 and various deletions of NS1 were cloned in frame with the GAL4DB and cotransfected into COS-7 and LA9 cells with a synthetic GAL4-responsive reporter plasmid. These studies showed NS1 can directly activate transcription through its 129 carboxyl-terminal amino acid residues. Any deletion from this region of the C terminus, even as few as 8 amino acids, completely abolishes transactivation. A yeast two-hybrid system used to identify protein-protein interactions demonstrated that NS1 is able to dimerize when expressed in yeast cells. However, only an almost complete NS11-638 bait was able to interact with the full-length NS1. A two-hybrid screen identified a HeLa cell cDNA clone (NS1-associated protein 1 [NSAP1]) that interacts with NS11-276 and NS11-638. An additional sequence was predicted from human EST (expressed sequence tag) data, and the cDNA was estimated to be at least 2,221 bp long, potentially encoding a 562-amino-acid protein product. A polyclonal antibody raised to a synthetic peptide within NSAP1 recognizes an approximately 65-kDa cellular protein. This NSAP1 cDNA has not previously been characterized, but the predicted protein sequence is 80% identical to the recently identified heterogeneous nuclear ribonucleoprotein (hnRNP) R (W. Hassfeld et al., Nucleic Acids Res. 26:439-445, 1998). NSAP1 contains four ribonucleoprotein domains, as well as a highly repetitive C-terminal region. A closely related mouse cDNA (deduced from murine EST data) encodes a protein with only a single amino acid residue change from the human protein. NSAP1 is predicted to be a 65-kDa polynucleotide binding protein, and it likely functions in the regulation of splicing and/or transport of mRNAs from the nucleus.

Megenteron manteri n. sp. and Steringophorus sp. (Digenea: Fellodistomidae) from Monomitopus agassizzi (Ophidiidae) in the Gulf of Mexico.

During a study of digenean parasites of deep-sea fishes, 81% (17 of 21) of Monomitopus agassizzi (Ophidiidae) from the Gulf of Mexico were infected with Megenteron manteri n. sp. and 10% (2 of 21) were infected with what appears to be a new species of Steringophorus. M. manteri n. sp. differs from M. crassum in having a longer body (3,733-9,394 microns), shorter caeca and a uterus that extends posteriorly beyond the ends of the caeca. The species of Steringophorus differs from other species in the genus in having smaller eggs (19-23 microns long) and caeca that extend into the posterior half of the hindbody.