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Background: This systematic review sought to characterize child-to-child and child-to-adult transmission of severe acute respiratory coronavirus 2 (SARS-CoV-2)., Methods: A systematic review was conducted from April 1, 2021, to December 15, 2023, to estimate secondary attack rates (SARs) and secondary infections per index case (case rate) from index cases up to age 20 years. SAR and case rate were analyzed based on age, setting, country and variant prevalence. Meta-analysis was conducted on the SAR data., Results: Eighty-six studies were included, representing 33,674 index cases. The total pooled SAR was 0.11 (95% CI: 0.07-0.16); 0.05 (95% CI: 0.03-0.10) for child-to-child transmission and 0.15 (95% CI: 0.07-0.30) for child-to-adult transmission. Pooled SAR in households was 0.28 (95% CI: 0.24-0.34) and was 0.02 (95% CI: 0.01-0.04) in schools., Conclusions: The role of children in SARS-CoV-2 transmission is small, particularly in schools. This work can help inform policies that effectively reduce transmission while minimizing adverse effects on children., Competing Interests: M.S. has been an investigator on projects funded by GlaxoSmithKline, Merck, Moderna, Pfizer and Sanofi-Pasteur. All funds have been paid to his institute, and he has not received any personal payments. L.S. has a leadership role with the Canadian Pediatric Society (which is unpaid). The other authors have no conflicts of interest to disclose., (Copyright © 2025 Wolters Kluwer Health, Inc. All rights reserved.)

Despite the availability of school-based immunization programs (SBIPs) in Canada, human papillomavirus (HPV) vaccine uptake remains suboptimal. Vaccine education may improve vaccine uptake among adolescents. The objective of this qualitative study was to identify opportunities for HPV vaccine education in British Columbia, Canada, by exploring the perspectives of students, parents, school staff, and public health nurses on the current SBIP. Individual semi-structured interviews were conducted with adult participants and focus groups were conducted with grade 6 students between November 2019 and May 2020. The interviews and focus groups were transcribed and then analyzed using reflexive thematic analysis. Opportunities for HPV vaccine education were identified in three themes: 1) making SBIPs student-centered; 2) adopting a collaborative and interdisciplinary approach to vaccine education; and 3) actualizing parent education opportunities. Broad support existed for a formal, collaborative HPV grade 6 vaccine curriculum delivered by teachers and public health nurses to provide evidence-based health information. Participants voiced that the curriculum should integrate students' perspectives on topics of interest and address needle associated pain and anxiety. Parents were identified as the primary vaccine decisionmakers, therefore, participants stated it was crucial to also provide parent-directed vaccine education as part of SBIP. Our findings support the development of a collaborative HPV vaccine curriculum directed to and informed by students and parents to buttress current SBIPs in British Columbia.

Background: Men who have sex with men (MSM) are at risk for sexually-transmitted hepatitis C (HCV). Evidence for HCV infection in the context of pre-exposure prophylaxis (PrEP) use in North America is limited. We sought to characterize baseline HCV prevalence and incidence in MSM receiving PrEP in British Columbia (BC), Canada., Methods: We followed individuals in the BC PrEP program from January 2018 to August 2019. We evaluated baseline prevalence and incident seroconversions (newly positive HCV antibody). A multivariable logistic regression model was performed in MSM for factors associated with HCV prevalence at enrollment, including reported prior sexually transmitted infection (STI), HIV Incidence Risk Index for MSM score, PrEP use because of a partner living with HIV, and location of residence., Results: The median age of the cohort was 33 years, 98.3% male, with 3058 person years (PY) of follow-up. Baseline HCV prevalence was 0.82% (31/3907 MSM enrollees) and HCV incidence (n = 3) was 0.15 per 100 PY (95% confidence interval [CI] 0.03-0.45). In multivariable analysis, initiating PrEP because of a partner living with HIV (adjusted odds ratio [aOR] 5.02; 95% CI 1.87-13.47) and prior STI (aOR 2.34; 95% CI 1.04-5.24) were associated with positive HCV status., Conclusions: Baseline HCV prevalence and incidence was low amongst MSM in a population-based PrEP program in BC, Canada. HCV was associated with bridging from populations living with HIV and evidence of a reported prior STI as a PrEP indicator condition amongst MSM. PrEP initiation may be an opportunity for linkage to HCV screening and treatment., (© 2022 John Wiley & Sons A/S . Published by John Wiley & Sons Ltd.)

Background: Artificial intelligence (AI)-driven symptom checkers are available to millions of users globally and are advocated as a tool to deliver health care more efficiently. To achieve the promoted benefits of a symptom checker, laypeople must trust and subsequently follow its instructions. In AI, explanations are seen as a tool to communicate the rationale behind black-box decisions to encourage trust and adoption. However, the effectiveness of the types of explanations used in AI-driven symptom checkers has not yet been studied. Explanations can follow many forms, including why-explanations and how-explanations. Social theories suggest that why-explanations are better at communicating knowledge and cultivating trust among laypeople., Objective: The aim of this study is to ascertain whether explanations provided by a symptom checker affect explanatory trust among laypeople and whether this trust is impacted by their existing knowledge of disease., Methods: A cross-sectional survey of 750 healthy participants was conducted. The participants were shown a video of a chatbot simulation that resulted in the diagnosis of either a migraine or temporal arteritis, chosen for their differing levels of epidemiological prevalence. These diagnoses were accompanied by one of four types of explanations. Each explanation type was selected either because of its current use in symptom checkers or because it was informed by theories of contrastive explanation. Exploratory factor analysis of participants' responses followed by comparison-of-means tests were used to evaluate group differences in trust., Results: Depending on the treatment group, two or three variables were generated, reflecting the prior knowledge and subsequent mental model that the participants held. When varying explanation type by disease, migraine was found to be nonsignificant (P=.65) and temporal arteritis, marginally significant (P=.09). Varying disease by explanation type resulted in statistical significance for input influence (P=.001), social proof (P=.049), and no explanation (P=.006), with counterfactual explanation (P=.053). The results suggest that trust in explanations is significantly affected by the disease being explained. When laypeople have existing knowledge of a disease, explanations have little impact on trust. Where the need for information is greater, different explanation types engender significantly different levels of trust. These results indicate that to be successful, symptom checkers need to tailor explanations to each user's specific question and discount the diseases that they may also be aware of., Conclusions: System builders developing explanations for symptom-checking apps should consider the recipient's knowledge of a disease and tailor explanations to each user's specific need. Effort should be placed on generating explanations that are personalized to each user of a symptom checker to fully discount the diseases that they may be aware of and to close their information gap., (©Claire Woodcock, Brent Mittelstadt, Dan Busbridge, Grant Blank. Originally published in the Journal of Medical Internet Research (https://www.jmir.org), 03.11.2021.)

Initiation of human immunodeficiency virus preexposure prophylaxis (PrEP) medications will also treat hepatitis B infection (HBV). The prevalence of chronic HBV was 0.86% (n=41/4760) among enrollees in a provincial PrEP program in British Columbia, Canada. Overall, 46.3% lacked follow-up HBV DNA monitoring, underscoring the need for HBV-related education for PrEP prescribers., (© The Author(s) 2021. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

GVHD, both acute and chronic, remains the major cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation. Thus, there is still a great need for therapeutic tools for the prevention and treatment of GVHD. Several biologics have shown promising results in salvage therapies but are attendant on an increased risk for opportunistic infections, lymphoproliferative disorders, and relapse. This is partly due to efficient T cell elimination that neither dissects alloreactive from non-alloreactive T cells nor considers functional and structural distinctiveness of pathogen- and malignancy-reactive γδ and iNKT T cells. A novel, humanized monoclonal antibody, GZ-αβTCR, specific for the human αβ T cell receptor, was evaluated in a xenogeneic GVHD model for its potential to prevent or ameliorate GVHD and prolong survival. We could show that GZ-αβTCR significantly attenuated clinical signs of GVHD and prolonged survival by preferential depletion of CD4 cells and the naïve T cell compartment, the trigger and driver of GVHD. In a regimen that included a preemptive dose, GZ-αβTCR treatment sufficiently abrogated GVHD. Importantly, GZ-αβTCR's specificity spared host cell-mediated immune competence of cell types other than αβT cells: namely γδT cells. GZ-αβTCR's outstanding capacity to prevent GVHD and ameliorate an ongoing GVHD while sparing immune cells other than αβT cells strongly recommends GZ-αβTCR for the prevention and treatment of acute GVHD in clinical settings.

Chronic graft-versus-host disease (cGVHD) is a significant hurdle to long-term hematopoietic stem-cell transplantation success. Insights into the pathogenesis and mechanistical investigations of novel therapeutic strategies are limited as appropriate animal models are missing. The immunodeficient NSG mouse - when humanized with human bone marrow, fetal liver and thymus (BLT NSG) - is prone for cGVHD, yet mainly affects the skin. In contrast, the NSG mouse humanized exclusively with CD34(+)-selected, CD3(+)-depleted stem cells (CD34(+)NSG) has neither been described for acute nor chronic GVHD so far. This is the first report about the development of systemic autoimmune cGVHD ≥24 weeks post stem cell receipt involving lung, liver, skin, gingiva and intestine in two NSG cohorts humanized with CD34(+) grafts from different donors. Affected mice presented with sclerodermatous skin, fibrotic lung, severe hepatitis, and massive dental malformation/loss. CD4(+)-dominated, TH2-biased, bulky T-cell infiltrates featured highly skewed T cell receptor (TCR) repertoires, clonal expansions, and autoreactive TCRs. In affected tissues profibrotic IL-13 and -4 dominated over TH1 cytokines IFN-γ and TNF-α. Thus, the time point of manifestation and the phenotype match human systemic pleiotropic sclerodermatous GVHD. The CD34(+)NSG-model's intrinsic deficiency of thymus, thymus-derived regulatory T cells (nTreg) and B cells emphasizes the role of the genetic polymorphism and the cytokines in the pathogenesis of cGVHD. Importantly, the only factor discriminating diseased versus non-diseased CD34(+)NSG cohorts were two risk HLA haplotypes that in human mediate susceptibility for autoimmune disease (psoriasis). Thus, the CD34(+)NSG model may serve as a platform for addressing issues related to the pathophysiology and treatment of human autoimmunity and chronic GVHD., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Stimulating the immune system to attack cancer is a promising approach, even for the control of advanced cancers. Several cytokines that promote interferon-γ-dominated immune responses show antitumor activity, with interleukin 12 (IL-12) being of major importance. Here, we used an antibody-IL-12 fusion protein (NHS-IL12) that binds histones of necrotic cells to treat human sarcoma in humanized mice. Following sarcoma engraftment, NHS-IL12 therapy was combined with either engineered IL-7 (FcIL-7) or IL-2 (IL-2MAB602) for continuous cytokine bioavailability. NHS-IL12 strongly induced innate and adaptive antitumor immunity when combined with IL-7 or IL-2. NHS-IL12 therapy significantly improved survival of sarcoma-bearing mice and caused long-term remissions when combined with IL-2. NHS-IL12 induced pronounced cancer cell senescence, as documented by strong expression of senescence-associated p16 INK4a and nuclear translocation of p-HP1γ, and permanent arrest of cancer cell proliferation. In addition, this cancer immunotherapy initiated the induction of myogenic differentiation, further promoting the hypothesis that efficient antitumor immunity includes mechanisms different from cytotoxicity for efficient cancer control in vivo .

Allograft rejection and immunosuppression are two major issues in transplantation medicine. The specific targeting of alloreactive T cells, the initiators and promoters of allograft rejection, would be a promising strategy to reduce unwanted T-cell responses and side effects of lifelong immunosuppression. The novel humanized monoclonal antibody GZ-αβTCR, specific for the human αβT-cell receptor, was tested in vitro and in vivo for its specificity and efficacy to modulate the αβT-cell compartment. GZ-αβTCR moderately induced apoptosis in resting αβT cells in vitro, an effect considerably amplified in activated T cells. A single dose of GZ-αβTCR significantly reduced human CD45(+)CD3(+) T cells in vivo, with a preferential modulation of CD4(+) αβT cells. Importantly, naive T cells, the T-cell subset from which alloreactivity emanates, were significantly reduced. Simultaneously, a significant, compensatory increase of γδ T cells was observed in vitro and in vivo in both humanized mouse models examined. GZ-αβTCR did not induce cytokines and was well tolerated. Thus, specificity and high efficacy make GZ-αβTCR a powerful tool to selectively eliminate putatively detrimental T-cell subsets, a major goal in transplantation medicine. At the same time, GZ-αβTCR spares γδ and natural killer cells, thus leaving the recipient's immune system competent for cell-mediated immunoregulation and cell-mediated immunity.

Background: Acute appendicitis frequently needs acute surgical intervention. Laparoscopic appendectomy (LA) and conventional open appendectomy (OA) are well established procedures, but appendectomy for intraoperative inconspicuous or vascular injected appendixes remains under debate because of potential postoperative morbidity. The aim of this study was to correlate intraoperative nonacute appendixes with histological and clinical outcome., Methods: Between 2005 and 2009, a total of 1,017 patients underwent OA or LA. A total of 1,005 patients were enrolled with inclusion criteria of suspicious acute appendicitis preoperatively. One hundred twenty-nine of 1,005 patients had intraoperative normal, vascular injected or chronic appendixes that were defined as nonacute appendicitis. Intraoperative findings were correlated with histological results and clinical outcome of patients., Results: Of 129 (12.8 %) of 1,005 patients with macroscopically nonacute appendicitis intraoperatively, 16.3 % had normal findings, 81.4 % vascular injection, and 2.3 % chronic alterations; and 94.6 % of nonacute appendixes had histopathological alterations: 38.9 % chronic, 14.0 % neurogenic, 26.4 % acute, 13.2 % phlegmonous, and 2.3 % malignant. Coproliths were found in 21.7 % of patients, most in vascular injected appendixes. Four of seven patients with histopathological normal appendixes had coproliths. Morbidity rate was 2.3 %, with no mortality., Conclusions: Appendiceal resection for intraoperative nonacute appendixes should be recommended because of high incidence of histopathological findings with low morbidity. In particular, chronic and neurogenous alterations cannot be predicted clinically or verified by radiological examination, but may cause recurrent symptoms.

Acute graft-versus-host disease is uncommon after liver transplant. We recently treated a 60-year-old man with liver transplant for hepatocellular carcinoma. After the primary liver transplant graft did not function, revision liver transplant resulted in excellent function. Subsequently, the patient developed watery diarrhea, systemic inflammatory response syndrome, a skin rash on his limbs and trunk, and palmar erythema. Skin biopsy suggested viral exanthems consistent with cytomegalovirus. Despite treatment for cytomegalovirus, intestinal symptoms worsened. Analysis of peripheral blood with fluorescence-activated cell sorting showed a high proportion of T lymphocytes, with 5% to 10% T cells specific to the second donor, suggestive of graft-versus-host disease. Within 48 hours after beginning therapy with antibodies against tumor necrosis factor-α (infliximab), the skin rash disappeared and endoscopy showed slight improvement of the mucosal regeneration. However, despite antifungal prophylaxis with caspofungin, the patient developed angioinvasive pulmonary aspergillosis and multiple organ failure, and he died. In conclusion, typical clinical symptoms of graft-versus-host disease after liver transplant may include skin rash and gastrointestinal symptoms, and diagnosis may be confirmed by histologic examination and testing for blood chimerism. A consensus for the treatment of graft-versus-host disease still is lacking, but tumor necrosis factor-α is an encouraging target for therapy to decrease the symptoms of graft-versus-host disease and enable mucosal regeneration.

Enteric duplications are rare, but can occur anywhere along the digestive tract. Most of the patients become symptomatic in early childhood and only a few cases of adult patients have been reported in literature. Here we report a unique case of an adenocarcinoma arising in a coincidentally found cystic duplication of the small bowel.

Quantification of virus removal by the purification process during production is required for clinical use of biopharmaceuticals. The current validation approach for virus removal by chromatography steps typically involves time-consuming spiking experiments with expensive model viruses at bench scale. Here we propose a novel, alternative approach that can be applied in at least one instance: evaluating retroviral clearance by protein A chromatography. Our strategy uses a quantitative PCR (Q-PCR) assay that quantifies the endogenous type C retrovirus-like particle genomes directly in production Chinese Hamster Ovary (CHO) cell culture harvests and protein A pools. This eliminates the need to perform spiking with model viruses, and measures the real virus from the process. Using this new approach, clearance values were obtained that was comparable to those from the old model-virus spike/removal approach. We tested the concept of design space for CHO retrovirus removal using samples from a protein A characterization study, where a wide range of chromatographic operating conditions were challenged, including load density, flow rate, wash, pooling, temperature, and resin life cycles. Little impact of these variables on CHO retrovirus clearance was found, arguing for implementation of the design space approach for viral clearance to support operational ranges and manufacturing excursions. The viral clearance results from Q-PCR were confirmed by an orthogonal quantitative product-enhanced reverse transcriptase (Q-PERT) assay that quantifies CHO retrovirus by their reverse transcriptase (RT) enzyme activity. Overall, our results demonstrate that protein A chromatography is a robust retrovirus removal step and CHO retrovirus removal can be directly measured at large scale using Q-PCR assays.

The effects of diet type (corn- vs. wheat-based) and multicarbohydrase addition on growth performance, digesta pH and viscosity, intestinal populations of Clostridium perfringens and lactic acid bacteria, and gut lesion score (from 0 to 4, where 0 = no gross lesions, 4 = severe extensive necrosis) of broiler chickens during oral challenge with C. perfringens (none or 10(8) cfu/bird on d 13) were studied in a 39-d experiment. A total of 1,216 male Ross-308 chickens was assigned to 8 dietary treatments in a randomized complete block design providing 8 replicate pens per treatment. Diets were formulated to meet the NRC protein requirement but were suboptimal in energy level. When compared with birds fed corn-based diets, chickens fed wheat-based diets had inferior (P < 0.01) final BW (2.49 vs. 2.59 kg) and feed conversion ratio (FCR; 1.83 vs. 1.78). Pathogen challenge significantly (P < 0.05) impaired growth performance and increased C. perfringens numbers and average lesion score. Increased (P < 0.01) C. perfringens counts (2.4 vs. 1.5 log(10) cfu/g of digesta) and intestinal lesion score (0.9 vs. 0.4) were observed for challenged birds fed wheat-based diets. No difference in digesta pH and lactic acid bacteria numbers were found among the treatments. Enzyme addition to both the corn- and wheat-based diets increased bird final BW (2.57 vs. 2.51 kg; P < 0.01), decreased overall FCR (1.78 vs. 1.83; P < 0.01), and, in those consuming wheat-based diets, reduced digesta viscosity (from 4.1 to 2.7 mPa.s; P < 0.01). Enzyme supplementation assisted the challenged birds in maintaining their optimal growth performance by improving (P < 0.05) average daily gain (59.5 vs. 56.9 g) in those consuming corn-based diets and FCR (1.83 vs. 1.90) in those consuming wheat-based diets to values similar to those observed in control birds (59.7 g/d and 1.84, respectively). In conclusion, enzyme addition improved growth performance and mitigated the negative effects of C. perfringens challenge.

We studied the role of CD28 in T-cell biology and T cell-mediated pathology using a novel mouse anti-mouse CD28 antibody, E18, which recognizes an epitope close to the B7 binding site. In vitro, this antibody completely blocked binding of B7 molecules to CD28 expressed on mouse thymocytes but enhanced anti-CD3-induced proliferation of peripheral T cells. Injections of E18 monoclonal antibody into normal BALB/c mice in vivo, however, led to a reversible reduction in Treg cell frequencies among CD4(+) cells, both in the thymus and in secondary lymphoid organs, suggesting that E18 acted as an inhibitor of CD28 signaling under these conditions. Antagonistic activity of E18 in vivo was further implied by suppressed responses of conventional CD4(+) T cells to stimulation with the superantigen staphylococcal enterotoxin B and in a model of acute graft-versus-host disease. In contrast to healthy mice, intact monoclonal antibody E18, but not its nonstimulatory Fab fragment, increased the frequencies of Treg cells among CD4(+) T cells in these pro-inflammatory settings allowing for efficacious protection from acute graft-versus-host disease. Thus, the agonistic signal generated by conventional, ie, nonsuperagonistic, anti-CD28 antibodies is important for their immunotherapeutic potential in vivo.

Use of hog (Sus scrofa) manure as a fertilizer is a practical solution for waste re-utilization, however, it may serve as a vehicle for environmental and domestic animal contamination. Work was conducted to determine whether pathogens, naturally present in hog manure could be detected in cattle (Bos taurus) grazed on the manure-treated pasture, and whether forage contamination occurred. During two 3 mo summer trials manure was applied to yield < or = 124 kg available N per hectare in a single spring or split spring and fall application. Samples of hog manure, forage, soil, and cattle feces were analyzed for naturally occurring Salmonella, Yersinia enterocolitica, and Escherichia coli. To follow movement of Salmonella in the environment isolates were identified to serovar and serotyped. Transfer of E. coli from hog manure to soil and cattle was examined by randomly amplified polymorphic DNA (RAPD) analysis of >600 E. coli isolates. While Y. enterocolitica was absent from all samples, in both years S. enterica Derby and S. enterica Krefeld were found in most hog manure samples, but were only on forage samples in the second year. Salmonella enterica Typhimurium, absent from hog manure was present on some forage in the first year. Cattle feces and soil samples were consistently Salmonella negative. These contaminations could not be traced to manure application. During this study, Salmonella and E. coli found in hog manure had different RAPD genomic profiles from those found in the feces of cattle grazing on manure-treated pasture.

A 21-day study was conducted to determine whether isoleucine might limit the performance of piglets fed low-crude protein (CP), amino acid (AA)-supplemented diets and to investigate the potential benefits of low-CP diets on gastrointestinal health in weaned pigs. Ninety-six piglets (initial BW = 6.44 ± 0.14 kg), housed four per pen, were randomly assigned to one of four diets, resulting in six replicate pens per diet. Dietary treatments were as follows: (1) 210 g/kg CP diet, (2) 190 g/kg CP diet deficient in isoleucine, (3) 190 g/kg CP diet supplemented with crystalline isoleucine up to the level in the 210 g/kg CP diet and (4) 170 g/kg CP diet supplemented with isoleucine and valine on the ideal protein ratio basis (60% and 70% relative to lysine, respectively). Pigs were allowed to adapt to the new environment for 4 days before the experiment commenced. Overall, pigs fed the 210 g/kg CP diet had higher (P < 0.05) average daily gain and lower (P < 0.05) feed : gain ratio compared with those fed the other diets. The faecal consistency score of pigs fed the 210 g/kg CP diet was higher (P < 0.05) than those fed the other diets. Pigs fed the 170 g/kg diet had lower (P=0.02) small intestine weight than those fed the 210 g/kg CP diet. Pigs fed the 210 g/kg CP diet had deeper (P < 0.05) crypt in the duodenum and ileum and higher (P < 0.05) ammonia N concentration in caecal digesta than those fed the other diets. There were no effects of diet on microbial population and volatile fatty acid concentration in the caecal digesta except for propionic acid whose concentration was higher (P < 0.05) for pigs fed the 170 g/kg diet than those fed the 190+isoleucine and the 210 g/kg CP diets. The results indicate that the low-CP, AA-supplemented diet reduced crypt hypertrophy, ammonia N concentration in the caecal digesta, small intestine weight and the performance of piglets. Also, the results of the current study were inconclusive with respect to whether isoleucine may limit the performance of pigs fed a low-CP, AA-supplemented diet.

The robustness of virus clearance with respect to protein A media reuse was demonstrated using media with four matrix chemistries: Protein A immobilized ProSep A, Poros A50, Protein A ceramic Hyper DF and MabSelect SuRe, an alkali resistant protein A ligand. Endogenous retrovirus clearance, step yield, impurity clearance and other performance parameters were evaluated periodically in media cycled up to 300 times. Media lifetime was generally limited by either declining step yield or media fouling. However, clearance of endogenous retrovirus remained in an acceptable range, either increasing or remaining constant. Multiply cycled media were tested for clearance of three viruses (SV40, X-MuLV, and MMV); clearance was comparable to naïve media. Overall, virus clearance by protein A chromatography appears to be extremely robust with respect to media age.

Phytosterols play a major role in functional foods. Their ability to reduce serum cholesterol in humans has been widely proven and they are now being added to various dairy based products. The present study investigated the potential antimicrobial activity of phytosterols in milk and their effect on yogurt starter cultures. A commercial phytosterol (0.26% to 1.8%, w/v) preparation (CPP) consisting of ss-sitosterol, campesterol, sitostanol, and campestanol had no effect on the standard plate count (SPC) and psychrotroph population in pasteurized milk stored at 4 degrees C. In addition, a challenge study employing Pseudomonas spp. in milk at 4 to 7 degrees C confirmed that the CPP was not antimicrobial. However, the addition of a dispersible CPP consisting of 0.72% phytosterol containing 0.02% to 0.03% sodium stearoyl lactylate (SSL) did appear to affect the SPC and psychrotrophic bacteria in refrigerated milk. The dispersible preparations did not, however, inhibit the growth of Pseudomonas. An investigation into the antimicrobial activity of SSL revealed that it alone had no effect on the SPC in milk. The CPP had no effect on growth and acid development by Lactobacillus bulgaricus and Streptococcus thermophilus during yogurt production at 33 degrees C and storage at 4 degrees C for 30 d. This is seen as a beneficial feature since growth and acid development by these organisms are crucial for yogurt quality. Saccaromyces cerevisiae and Aspergillus ochraceous added to yogurt as typical contaminants also were not inhibited. While the CCP was somewhat antimicrobial when formulated with dispersing agents, it otherwise had no antimicrobial activity.

Aims: To examine the effect of processing with superheated steam (SS) on Geobacillus stearothermophilus ATCC 10149 spores., Methods and Results: Two inoculum levels of spores of G. stearothermophilus were mixed with sterile sand and exposed to SS at 105-175 degrees C. The decimal reduction time (D-value) and the thermal resistance constant (z-value) were calculated. The effect of cooling of spores between periods of exposure to SS was also examined. A mean z-value of 25.4 degrees C was calculated for both inoculum levels for SS processing temperatures between 130 degrees C and 175 degrees C., Conclusions: Spore response to SS treatment depends on inoculum size. SS treatment may be effective for reduction in viability of thermally resistant bacterial spores provided treatments are separated by intermittent cooling periods., Significance and Impact of the Study: There is a need for technologies that require short thermal processing times to eliminate bacterial spores in foods. The SS processing technique has the potential to reduce microbial load and to modify food texture with less energy in comparison to commonly used hot air treatment. This work provides information on the effect of SS processing parameters on the viability of G. stearothermophilus spores.

The effects of ground flaxseed (FS) and a multicarbohydrase enzyme (C) supplement on piglet performance, gastrointestinal microbial activity, and nutrient digestibility were investigated in a 28-d trial. The enzyme supplement provided 500 units of pectinase, 50 units of cellulase, 400 units of mannanase, 1,200 units of xylanase, 450 units of glucanase, and 45 units of galactanase per kilogram of diet. Ninety-six pigs were weaned at 17 d of age (BW, 6.1 +/- 0.4 kg, mean +/- SD) and assigned to treatments based on a 2 x 2 factorial arrangement in a completely randomized design, with 6 pens per diet (4 pigs per pen). The diets contained wheat, barley, peas, soybean meal, and canola meal with 0 or 12% FS, and were fed without or with C. Flaxseed was included by changing the levels of the other ingredients to balance the diets for DE and nutrients. Diets had similar nutrient contents and met the NRC (1998) nutrient specifications, with the exception of DE, CP, and AA, which were 95, 94, and 97% of the NRC requirements, respectively. Diets were fed in a 2-phase feeding program (2 wk/phase). Feed intake and BW were measured weekly, and 1 pig per pen with a BW nearest the pen average was bled weekly to evaluate plasma urea nitrogen. On d 28, fresh fecal samples were collected from each pen and 1 pig per pen with a BW nearest the pen average was killed to evaluate intestinal microbial activity and nutrient digestibility. A dietary effect on piglet performance was observed only in wk 3, when the FS diets decreased (P = 0.005) ADG and G:F, tended to decrease (P = 0.070) ADFI, and increased (P = 0.027) plasma urea nitrogen. An interaction between FS and C was observed for ileal digesta viscosity (P = 0.045), such that C increased viscosity in the FS diet but had no effect in the non-FS diet. Flaxseed and C interacted to affect ileal ammonia content (P = 0.049), such that in the absence of FS, pigs fed the diet with C had lower ammonia than those on the diet without C. Flaxseed and C affected other ileal parameters independently. Pigs fed the FS diets had decreased (P = 0.003 to 0.033) anaerobic spore counts, organic acid, DM, CP, and nonstarch polysaccharide (NSP) digestibility compared with pigs fed the non-FS diets, whereas pigs fed the C-supplemented diets had greater (P = 0.009 to 0.061) lactobacilli counts, lactate, DM, and NSP digestibility than pigs fed the unsupplemented diets. In conclusion, FS reduced ileal microbial activity, nutrient digestibilities, and piglet performance in wk 3. The multicarbohydrase supplement increased ileal DM and NSP digestibilities as well as lactobacilli counts and lactate.

The effect of incorporating trisodium phosphate (TSP) in pea starch (PS) and acidified sodium chlorite (ASC) in calcium alginate upon the antimicrobial activity of TSP and ASC was studied against a 3-strain cocktail of Salmonella inoculated on chicken skin. The influence of polymer coating concentration on skin pH, coating-skin adhesion, and coating absorption upon antimicrobial performance were investigated. Aqueous solutions of 0.5 to 4.8% (wt/vol) PS were prepared with 10% (wt/vol) TSP (PS + TSP coating), and alginate + ASC coatings contained 1% (wt/vol) calcium chloride in 1,200 ppm of ASC mixed with an aqueous solution of 0.5, 1.0, or 1.5% (wt/vol) sodium alginate. Coating drops (10 microL) were placed on chicken skin thighs, and the angle formed by the tangent of the liquid surface at the skin interface (contact angle) was measured using a digital camera to assess coating-skin adhesion. Excised skins were mounted in a ring holder, and 5 mL of the coatings was applied to the skin. Weight changes in the skins that were related to coating absorptiveness were recorded. The TSP dissolved in 3.5% PS and ASC in 1% alginate reduced Salmonella by 1.6 log cfu/g and 1.4 log cfu/g, respectively, within 24 h. These reductions were significantly greater than those caused by TSP or ASC alone in water for up to 120 h. In coatings, TSP and ASC caused significant elevation or reduction of skin surface pH for up to 120 h, respectively. The TSP destabilized PS with 88% of the coating having dripped from the skin 1 h later. Coatings with 0.5% PS were absorbed quickly by the skin and had high skin adhesion, whereas those with >3.5% PS had low skin adhesion and slow absorption. Alginate coatings with or without ASC were stable, and about 50% of the coating weight was retained at 120 h. The latter coatings appeared to have low absorptiveness because the skin gained approximately 1.0% of its weight within 60 min following application. These findings indicate that effects of the agents in coatings on skin pH, the extent of coating adhesion, and absorption may contribute to overall antimicrobial behaviors.

Addition of animal manure to soil can provide opportunity for Salmonella contamination of soil, water, and food. This study examined how exposure of hog manure-treated loamy sand and clay soils to different simulated seasonal temperature sequences influenced the length of Salmonella survival. A six-strain cocktail of Salmonella serovars (Agona, Hadar, Heidelberg, Montevideo, Oranienburg, and Typhimurium) was added to yield 5 log cfu/g directly to about 5 kg of the two soils and moisture adjusted to 60 or 80% of field capacity (FC). Similarly, the Salmonella cocktail was mixed with fresh manure slurry from a hog nursery barn and the latter added to the two soils at 25 g/kg to achieve 5 log cfu/g Salmonella. Manure was mixed either throughout the soil or with the top kilogram of soil and the entire soil volume was adjusted to 60 or 80% FC. Soil treatments were stored 180 d at temperature sequences representing winter to summer (-18, 4, 10, 25 degrees C), spring to summer (4, 10, 25, 30 degrees C), or summer to winter (25, 10, 4, -18 degrees C) seasonal periods with each temperature step lasting 45 d. Samples for Salmonella recovery by direct plating or enrichment were taken at 0, 7, and 15 d post-inoculation and thereafter at 15-d intervals to 180 d. Salmonella numbers decreased during application to soil and the largest decreases occurred within the first week. Higher soil moisture, manure addition, and storage in the clay soil increased Salmonella survival. Salmonella survived longest (> or = 180 d) in both soils during summer-winter exposure but was not isolated after 160 d from loamy sand soil exposed to other seasonal treatments. For all but one treatment decimal reduction time (DRT45d) values calculated from the first 45 d after application were < or = 30 d and suggested that a 30-d delay between field application of manure in the spring or fall and use of the land would provide reasonable assurance that crop and animal contamination by Salmonella would be minimized.

The effects of low-protein AA-supplemented diets on piglet performance, visceral organ mass, incidence of diarrhea, intestinal microbial population, and fermentation were studied in a 3-wk trial. After a 7-d adaptation period, 96 piglets (approximately 6.2 kg of initial BW) were assigned to 4 corn-wheat, soybean meal-based dietary treatments in a completely randomized design to give 6 replicate pens per treatment (n = 4 piglets per pen). The treatments were a control wheat-corn-soybean meal-based phase I diet containing 23% CP, or the same diet with CP reduced to 21%, 19%, or 17% and supplemented with crystalline AA to achieve equal standardized ileal digestible contents of Lys, Met plus Cys, Thr, and Trp in all diets. Diets were formulated to similar nutrient levels and provided ad libitum. Blood from all pigs was taken on d 0, 7, 14, and 21 for determining plasma urea N. Weekly feed intake, BW changes, and G:F were determined. On d 21, 2 pigs per pen were randomly selected and killed to determine small intestinal morphology, digesta pH and ammonia levels, and luminal microbial counts. Average daily feed intake, ADG, and G:F were not affected (P > 0.10) by reducing CP to 21%, but a reduction to 19% or 17% decreased ADFI (P < 0.001) and ADG (linear, P < 0.001; quadratic, P < 0.05) over the 3-wk study period. Reducing CP to 19% had no effect (P > 0.10) on G:F; however, this response criterion was decreased linearly (P < 0.001) over the 3-wk study period as dietary CP declined. Water usage was only numerically decreased (P > 0.10) with dietary CP reduction. Plasma urea N was decreased linearly (P < 0.01) with CP reduction. Reducing CP from 23 to 17% had a linear (P < 0.05) and cubic effect on stomach and liver weights, respectively. Although histological data showed some differences among diets, no distinct trend was evident. Ammonia N in ileal digesta was reduced linearly (P < 0.01) as dietary CP was decreased. With the exception of valeric acid, VFA levels in ileal digesta of piglets fed low-protein diets were generally lower (P < 0.05) compared with the control diet. Diet had no effect on intestinal microbial counts (P > 0.10). The results show that piglet performance may suffer when dietary CP is reduced by 4 or more percentage units from 23% and support the hypothesis that low-CP diets help maintain enteric health in pigs by lowering toxic microbial metabolites such as ammonia.

Background: Hyperoxaluria has been incriminated to account for the increased incidence of urolithiasis or nephrocalcinosis in patients with cystic fibrosis (CF). Hyperoxaluria presumably is caused by fat malabsorption and the absence of such intestinal oxalate-degrading bacteria as Oxalobacter formigenes. To better elucidate its pathophysiological characteristics, we prospectively studied patients with CF by determining these parameters and performing renal ultrasonography twice yearly., Methods: In addition to routine tests in urine (lithogenic and stone-inhibitory substances), the presence of O formigenes was tested in stool, plasma oxalate was measured, and a [13C2]oxalate absorption test was performed in 37 patients with CF aged 5 to 37 years (15 females, 22 males) who were constantly hyperoxaluric before the study., Results: Hyperoxaluria (oxalate, 46 to 141 mg/1.73 m2/24 h [0.51 to 1.57 mmol/1.73 m2/24 h]; normal, < 45 mg/1.73 m2/24 h [< 0.5 mmol/1.73 m2/24 h]) was now found in 24 patients (64.8%). Plasma oxalate levels were elevated in 6 patients (7.92 to 19.5 micromol/L; normal, 6.3 +/- 1.1 micromol/L). Oxalobacter species were detected in only 1 patient. Intestinal oxalate absorption was elevated (11.4% to 28.5%; normal, < 10%) in 23 patients. Hypocitraturia was present in 17 patients (citrate, 0.35 to 2.8 g/1.73 m2/24 h [0.2 to 1.1 mmol/1.73 m2/24 h]; normal female, > 2.8 mg/1.73 m2/24 h [> 1.6 mmol/1.73 m2/24 h]; male, > 3.3 mg/1.73 m2/24 h [> 1.9 mmol/1.73 m2/24 h]). Urine calcium oxalate saturation was elevated in 17 patients (5.62 to 28.9 relative units; normal female, < 5.5 relative units; male, < 6.3 relative units). In 16% of patients, urolithiasis (n = 2) or nephrocalcinosis (n = 4) was diagnosed ultrasonographically., Conclusion: Absorptive hyperoxaluria and hypocitraturia are the main culprits for the increased incidence of urolithiasis and nephrocalcinosis in patients with CF. We advocate high fluid intake, low-oxalate/high-calcium diet, and alkali citrate medication, if necessary. Additional studies are necessary to determine the influence of Oxalobacter species or other oxalate-degrading bacteria on oxalate handling in patients with CF.

Harvesting of Brazil nuts not only helps to preserve the Amazon rainforest but also provides income to individuals who would otherwise have little means of making a livelihood. Recently, the European Community has tightened the quality requirements for Brazil nuts, particularly with regard to aflatoxin levels and microbiological contamination. The objectives of this research were to gain a better understanding of the origin of aflatoxins on Brazil nuts and to microbiologically evaluate some of the operations involved in processing. In this regard, five Brazil nut pods were aseptically picked from trees located in each of three concessions of the Peruvian Amazon rainforest (Madre de Dios province). The exteriors of the pods and the nuts were examined for yeast and molds, including Aspergillus flavus and Aspergillus parasiticus, and for bacteria, including Salmonella and Escherichia coli. Brazil nuts obtained from various commercial process operations located in Peru were similarly evaluated. Exteriors of all Brazil nut pods did not contain A. parasiticus, and only pods from one concession yielded A. flavus isolates. All isolates tested were aflatoxigenic (630 to 915 ppb total aflatoxin). Coliforms, E. coli, and salmonellae were not recovered from any of the pods. Whole, in-shell nuts obtained after opening the pods yielded no A. flavus or A. parasiticus. Aflatoxins were not detected (detection limit 1.75 ppb) in any of the nuts. Whole, in-shell and shelled nuts from various process operations were all positive for A. flavus but negative for E. coli and salmonellae. Soaking of whole, in-shell nuts before cracking or shelling increased coliform numbers, whereas levels of A. flavus decreased. In order to gain a better understanding of the sanitary performance of the unit process operations, additional evaluations should be conducted on product lots processed on different days. Also, the microbiology of product processed from common lots should be followed through the various unit operations and compared.

We evaluated viral clearance in multiply-cycled anion-exchange media run in flow-through mode. We found that anion-exchange columns do not lose viral clearance capacity after extensive re-use, if they are cleaned with recommended buffers that do not chemically degrade the media. In contrast, anion-exchange (AEX) columns that are not cleaned or are cleaned with buffers that chemically degrade the media lost viral clearance capacity after extended use. In these cases, other performance attributes that changed at the same time were increased band spreading, decreased DNA clearance and accumulating backpressure that prevented re-use past 80-120 cycles. Thus, our data suggests that flow through mode anion-exchange columns that are cleaned with recommended cleaning buffers, and periodically monitored for band spreading, DNA clearance and/or backpressure need not be re-evaluated for viral clearance at the end of the validated media lifetime.

The ability of Salmonella, Escherichia coli O157:H7, Listeria monocytogenes, and Shigella to survive or grow in pesticide solutions (Ambush 240EC, Benlate T-N-G, Bravo 500, Botran 75WP, Captan 80WDG, Parasol, and Vendex 50W) used by the horticultural industry was examined. In the laboratory, individual cultures were inoculated at 4 log CFU/ml in pesticides diluted with sterile saline to the lowest recommended spray concentrations. During 21 degrees C incubation for < or =96 h, bacterial survivors in the samples and a control consisting of saline were enumerated either by agar surface plating or hydrophobic grid membrane filtration. Most formulations tested were somewhat inhibitory to the pathogenic bacteria. All inoculated bacteria survived or grew in Bravo 500. Among bacteria tested, Salmonella spp. were best able to survive and Listeria spp. were least able to survive in pesticide solutions. When the incubation temperature or pesticide concentration was increased, survival of Salmonella varied depending on the type of formulation. In the field, when a bacterial cocktail containing E. coli O157:H7 and Salmonella Enteritidis was added to Bravo 500 at 6 log CFU/ml, both organisms were recovered from leaves and fruit skins of sprayed tomato plants after the recommended 1 day-to-harvest interval. E. coli and Salmonella survived longer on tomato leaves when sprayed in saline (at least 26 and 56 days, respectively) than when sprayed in Bravo 500 (>45 h and <15 days, respectively). While Salmonella serovars Typhimurium and Heidelberg grew in the fungicide Bravo, and Enteritidis grew in the insecticide Vendex within 96 h at 21 degrees C in the laboratory, pathogen growth in other pesticide formulations did not occur. Higher temperature (< or =30 degrees C) or doubling pesticide concentrations had either no or a negative effect on Salmonella Heidelberg survival. Use of unexpired pesticide formulations may have contributed to the reduced bacterial survival and growth found in the laboratory and during the field trials with Bravo.

The survival of Escherichia coli O157:H7 strains in both frozen foods and trypticase soy broth (TSB) was investigated following cold shocking at 10 degrees C for 1.5 h. Using both trypticase soy agar (TSA) and violet red bile agar (VRBA) as recovery media, it was demonstrated that survival levels between cold shocked (CS) and non-cold shocked (NS) E. coli in ground beef or pork were not significantly different (P < or = 0.05). In contrast, cold shocking E. coli in either milk, whole egg or sausage resulted in a significant(P < or = 0.05) enhancement in survival. For milk, survival levels of CS E. coli, by 28 days of frozen storage, were 1.89 and 1.66 log10 cfu/ml higher on TSA and VRBA, respectively, when compared to NS cells. In egg these values were 0.64 and 1.31, while in sausage, values of 0.74 and 1.19 were obtained. In TSB (pH 7) survival of CS E. coli for some strains was about 3 log10 cfu/ml higher when compared to NS cells. Acidification of TSB (pH 5), however, appeared to negate the protective effects of the cold shock treatment. In milk, increasing the differential between the growth and cold shock temperatures resulted in higher numbers of survivors. In this regard the growth-cold shock temperature protocol giving optimum protection was 37-10 degrees C. In contrast, growth of E. coli at 20 degrees C followed by cold shocking at 10 degrees C did not result in any significant freeze protection. In addition, increased protection due to cold shocking was correlated with the appearance of a novel protein appearing at pI 4.8 following isoelectric focusing analysis, thus demonstrating an alteration of protein synthesis.

The large-scale production of recombinant monoclonal antibodies demands economical purification processes with high throughputs. The potential for ion-exchange membrane adsorbers to replace traditional ion-exchange columns was evaluated. Breakthrough capacities of commercially available cation-exchange membranes were determined as a function of flow-rate and layer number. Due to economic and process restrictions, cation-exchange membranes may not currently be advantageous for process-scale antibody purification in a bind and elute mode. However, anion-exchange membranes in a flow-through mode may provide a reasonable alternative to columns for the removal of low levels of impurities such as DNA, host cell protein, and virus.

Expanded bed adsorption (EBA) was examined as the initial capture/purification step in the purification of monoclonal antibodies from Chinese hamster ovary (CHO) cultures. Two process alternatives each using EBA were compared to a conventional Protein A process without EBA. One alternative used Protein A affinity EBA followed by packed-bed cation and anion-exchange steps. The other alternative used cation-exchange EBA as the capture step followed by packed-bed Protein A and anion-exchange steps. The process using Protein A EBA produced comparable purity (host cell protein, DNA, Protein A, antibody aggregate) to the conventional process. However, the Protein A EBA column showed a significant decrease in dynamic capacity with a limited number of cycles. The process using cation EBA achieved comparable levels of host cell proteins (HCP) and DNA but not antibody aggregate or leached Protein A compared to the conventional process.

Three experiments were conducted to determine (1) the pharmacodynamics of 5-hydroxytryptamine in juvenile lobsters; (2) the effects of 5-hydroxytryptamine, using a range of dosages, on a motor behavior used to escape an aversive situation; and (3) the effect of doses that did and did not inhibit this motor behavior on measures of dominance and shelter competition. The fate of 5-hydroxytryptamine in hemolymph over a 60-min post-injection period showed that the concentration fell rapidly to a low plateau that was maintained for at least 1 h. Low doses of 5-hydroxytryptamine did not affect locomotor behavior, but higher doses inhibited it. Dominance and subsequent possession of a shelter were unaffected by a low dose of 5-hydroxytryptamine but a higher dose that inhibited locomotion resulted in lobsters that lost fights and did not secure or retain possession of the shelter. In the context of dominance and shelter competition, we were unable to demonstrate any advantage of the low dose of exogenous 5-hydroxytryptamine and a severe disadvantage with the higher dose. Previous reports of transient increases in aggression in 5-hydroxytryptamine-treated subordinate lobsters did not take into account motor inhibition as a possible critical variable in aggression.

We show that expanded bed protein A affinity chromatography using Streamline rProtein A media is an efficient method for purifying a recombinant humanized monoclonal antibody from unclarified Chinese hamster ovary cell culture fluid and that it provides purification performance comparable to using a packed bed. We determined that the dynamic capacity of the expanded bed media is related to flow rate (measured in column volumes per hour) by a power function, which allows a high capacity at a low flow rate. At 250 cm h-1 with a 25 cm bed height (10 column volumes h-1), the dynamic capacity is 30 g l-1. The yield and purity (measured by the amount of host cell proteins, DNA, SDS-PAGE, and turbidity) of the antibody purified by expanded bed is comparable to the yield and purity obtained on a standard packed bed method using Prosep A media.

We describe the performance characteristics of five Protein A affinity-chromatography sorbents (Sepharose Fast Flow, Poros 50, Poros LP, Prosep and Streamline) for purifying a recombinant humanized monoclonal antibody from clarified Chinese hamster ovary cell culture fluid. We measured the dynamic capacity at varying flow rates, maximum capacity, pressure drop and production rate. For purified antibody, we measured yield and purity (by SDS/PAGE, the amount of DNA, the amount of host-cell proteins and the amount of Protein A). We found that, whereas all sorbents provided significant and essentially equivalent antibody purification, there were differences in capacity and pressure drop, which affected the production rate and had implications for process applications.

We show that an on-line chromatographic assay can reliably control antibody loading in real-time during protein A affinity chromatography purification of a recombinant antibody from clarified Chinese hamster ovary cell culture fluid. The on-line assay directly sampled preparative column effluent and provided real-time measurement of antibody breakthrough during loading. The on-line assay used protein A immobilized on perfusion chromatography media, equilibrated with phosphate-buffered saline at pH 7.2 and eluted with phosphate-buffered saline at pH 2.2. The assay reliably ended loading at 1% breakthrough with minimal yield loss. Reproducible yield and purity were obtained over 23 consecutive cycles. Yield remained constant while breakthrough capacity and the antibody concentration in the load changed.

An on-line assay was developed to monitor antibody breakthrough in real time during Protein A affinity chromatography of recombinant antibodies. When loading cell culture fluid on to a Protein A affinity column, antibody breakthrough cannot be measured by UV absorbance because of the flow-through of UV-absorbing impurities. An assay using perfusion chromatography media with immobilized Protein A is a rapid, antibody-specific assay. It directly samples preparative column effluent, allowing real-time measurement of antibody breakthrough during loading of Protein A affinity chromatography. Breakthrough curves were generated for three column media at five flow rates, showing the effects of diffusion on the shape of the breakthrough curve. The breakthrough curves were used to measure dynamic capacity.

Acetonitrile is used as an eluent for reversed-phase chromatography. However, because it is a flammable solvent, using acetonitrile on a large scale requires expensive equipment and facilities specially designed for flammable solvents. Using a non-flammable solvent as an eluent eliminates this expense. A method was developed to purify recombinant human insulin-like growth factor I by reversed-phase high-performance liquid chromatography using gradient elution with hexylene glycol, a non-flammable replacement for acetonitrile. The separation produced equivalent yield, purity and throughput as reversed-phase chromatography using elution with acetonitrile.

During preparative reversed-phase chromatography of recombinant human insulin-like growth factor-I (IGF), the separation of IGF from IGF aggregates cannot be determined using UV absorbance. An on-line reversed-phase chromatographic assay was developed that provides a quantitative measurement of IGF and IGF aggregates every 4 min, allowing real-time control of purified IGF collection. Process control using the on-line assay is a reliable and accurate method to collect purified IGF.

A two-stage recovery protocol was examined for microorganisms following gamma irradiation in phosphate buffer at 0 degrees C. In the first stage, survivors were recovered on basal yeast extract agar and held at various temperatures suboptimal for their growth for 20 h (resuscitation protocol). In the second stage the survivors were incubated for an additional 24 h, but in this case at their optimum temperature for growth. Controls consisted of survivors which were not subjected to the resuscitation protocol (direct incubation at their optimum growth temperature). The ratio of survivors enumerated with and without the resuscitation protocol (control) at each specified temperature was used to formulate a recovery factor(RF). An RF was determined for each treatment dose. Results of this study indicated that the number of Escherichia coli, Salmonella serotype typhimurium and Brochothrix thermosphacta survivors increased following a resuscitation protocol (RF > 2.0). Overall, optimum resuscitation temperatures ranged from 14 to 22 degrees C. The extent of recovery also appeared dose dependent, with larger treatment doses giving rise to a higher RF. S. serotype typhimurium irradiated at 1.5 kGy exhibited the highest RF, 161, when resuscitated at 22 degrees C. Listeria monocytogenes, Yersinia enterocolitica, Staphylococcus aureus, Aeromonas hydrophila and Saccharomyces cerevisiae exhibited an RF < 2.0 regardless of resuscitation temperature. Results of this study indicated that the use of suboptimal holding temperatures as part of a recovery protocol may have advantages, especially with respect to the enumeration of E. coli and salmonellae survivors in irradiated foods such as poultry.

Male Mongolian gerbils (Meriones unguiculatus) 14-54 months old (n = 77) were evaluated in a battery of psychomotor (open field, locomotor, and runwheel activity, rotorod performance) and learning (one-way active avoidance in a straight runway and in 14-unit T-maze performance) tests. Body weight and seizure activity were also monitored. According to linear regression analysis, runwheel activity decreased with age; and the number of errors in the 14-unit T-maze increased as a function of age (ps < 0.05). None of the other behavioral measures or body weight were significantly correlated with age. This gerbil strain (Tumblebrook Farms; West Brookfield, MA) tended to be very prone to seizures with 64% of the gerbils experiencing at least one seizure while being tested. Seizures tended to occur when the gerbil was exposed to a novel situation (e.g., initial weighing, placement on the rotorod). An age-related decline in some aspects of psychomotor and learning performance was observed, suggesting the gerbil as an additional mammalian model of aging. The high incidence of seizure activity presented a complicating and confounding variable to the interpretation of the results of the behavioral tests used in the present study. Interventions to control seizure activity (e.g., systematic, controlled breeding; adaptation to apparati) in this model will likely increase its viability as a mammalian model of aging.

Purpose: Cardiopulmonary bypass is associated with activation of neutrophils, which may adhere to vascular endothelium causing lung, heart, and brain injury. We tested whether blocking neutrophil adherence would improve organ function following cardiopulmonary bypass in dogs., Materials and Methods: All dogs received a standard anesthetic, and then one group (n = 6) received 2 hours of cardiopulmonary bypass followed by 4 hours of observation. A second group (n = 6) received a monoclonal antibody (6 mg/kg) to CD18, a neutrophil adherence factor, immediately before cardiopulmonary bypass. A third group (n = 6) did not receive cardiopulmonary bypass or antibody., Results: Using flow cytometry we found that the antibody bound essentially all neutrophil CD18 sites. All three groups had similar gas exchange and hemodynamics. Lung and heart histology results were similar between groups. By echocardiography, five animals receiving cardiopulmonary bypass alone showed regional wall abnormalities, whereas only one receiving antibody showed wall motion abnormality (P < .05). Following cardiopulmonary bypass, intracellular myocardial pH was higher (P < .05) in the antibody-treated group compared with the group that had cardiopulmonary bypass alone (7.23 +/- 0.05 v 7.07 +/- 0.07 respectively)., Conclusion: Monoclonal antibodies to CD18 can prevent the deterioration in cardiac function routinely observed following cardiopulmonary bypass.

The use of microwave plasmas for dry sterilization has been investigated. The dry-sterilization process is a process similar to plasma etching. Bacteria and viruses can be killed by chemical reactions which disintegrate their bodies and remove them from the surface to be sterilized. The removal of bacteria or viruses from material surfaces is caused by the reaction of activated oxygen species in the plasma with hydrocarbon bonds of the cell wall of the bacteria or the capsid of the viruses. Preliminary experiments indicate that the low-temperature dry sterilization method is easy to use, requires much less time than other methods for sterilization, and is also non-toxic. It is feasible for use in the field of sterilization in dental and medical clinics.

Immersion chilling has been identified as a critical control point in commercial poultry processing. A study was undertaken to investigate the impact of immersion chilling on the microbiology of carcasses within a small to medium sized commercial operation. Fifty chilled carcasses (following immersion chilling) sampled over a 10-day period using a whole bird rinse technique exhibited mean standard plate count (SPC) and coliform counts (log colony-forming units (CFU)/ml) of 3.74 and 3.03, respectively. These levels were both significantly ( P < 0.005) lower (ca. 1 log unit) compared to similar numbers of prechill carcasses (birds exiting the inside-outside washer but prior to the prechiller). Water from the chiller was also shown to contain significantly ( P < 0.005) lower (ca. 1 log unit) SPC and coliform levels compared to those from the prechiller. Reducing the flow of water at the inside-outside washer by 50% did not significantly ( P < 0.001) affect the SPC and coliform levels of either prechilled or chilled carcasses.

The irradiation sensitivity of fungal spores to either gamma or electron beam irradiation was evaluated in distilled water. The D10 (the dose required to reduce the initial population by 90%) gamma values ranged from 0.236 to 0.416 kGy and from 0.209 to 0.319 kGy for Penicillium and Aspergillus species, respectively. The D10 electron beam values ranged from 0.194 to 0.341 and from 0.198 to 0.243 kGy for Penicillium and Aspergillus species, respectively. Of the aspergilli species evaluated, only half exhibited significantly (P < 0.05) greater sensitivity to the electron beam treatment compared to gamma irradiation. Four of the six penicillia species evaluated also exhibited significantly (P < 0.05) higher sensitivities to electron beam treatment.

There are three major methods for buffer exchange of proteins at industrial scale: size exclusion chromatography (SEC), tangential flow filtration (TFF), and countercurrent dialysis (CCD). In order to determine the optimal technology for a given process, a study was done to compare these technologies on a technological and economic basis. This comparison required that new mathematical models be developed which enable the common features of each unit operation to be directly compared. The new concept of a diavolume equivalent for SEC, defined as the inverse of the fractional loading, was also introduced to aid in this comparison. Variables that were examined for each unit operation included range of buffer exchange, dilution of protein solution, yield, buffer requirements, total operating time, throughput, plant space, capital, raw materials, and labor costs. It was found that TFF and CCD have a greater range of buffer exchange than SEC. TFF also provides the advantage that concentration of the protein can readily be accomplished in the same step. For processes of equal batch size and yield, TFF and CCD also provide a two- to five- fold improvement in each of the remaining variables. The major economic advantage in using TFF and CCD over SEC is the decreased plant size required for manufacturing and thus the longer term use of existing facilities. Situations where SEC (or CCD) would be favored over TFF are when protein denaturation occurs in TFF but does not occur in SEC. (c) 1995 John Wiley & Sons, Inc.

We previously developed an efficient route to humanized F(ab')2 fragments by high level secretion of the Fab' arms from Escherichia coli followed by directed chemical coupling in vitro. Here the number and type of interchain linkages in F(ab')2 molecules has been modified to simplify their production and improve their serum stability. All F(ab')2 variants had comparable binding affinity for the p185HER2 Ag and antiproliferative activity against p185HER2-overexpressing tumor cells. This was anticipated since the modifications are distant from the Ag-binding loops. Replacement of a single disulfide bridge between Fab' arms with a more stable thioether bridge increased the serum permanence time in normal mice by threefold to 2.1 h. Removal of the disulfide bond between L and H chains in the thioether-bridged F(ab')2 did not affect the pharmacokinetics, suggesting that the L chain remains associated with the H chain. An additional Fab' variant containing three repeats of the motif, CysProPro, was constructed with the aim of promoting efficient formation of F(ab')2 in E. coli. This Fab' (CPP)3 variant was recovered predominantly (up to 70%) as F(ab')2 directly from fermentation cell pastes, thus circumventing the need for in vitro coupling. The F(ab')2 (CPP)3 variant has a similar serum pharmacokinetics to the thioether-bridged molecules. The improvements described here for deriving F(ab')2 fragments from E. coli should enhance the clinical potential of these molecules.