Harold M. Barnhart’s research while affiliated with University of Nebraska–Lincoln and other places

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Publications (40)


Effect of γ-irradiation on bovine and human milk lysozyme
  • Article

June 2008

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10 Reads

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4 Citations

R. R. EITENMILLER

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H. M. BARNHART

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K. M. SHAHANI

SUMMARY –Lysozymes (mucopeptide N-acetylmuramylhydrolase, EC 3.2.1.17) isolated from human milk (HML) and bovine milk (BML) were subjected to -irradiation and their radiosensitivity was compared to that of egg white lysozyme (EWL). When irradiated in phosphate buffer, pH 6.2, the enzymes were fairly sensitive to -irradiation. Inactivation followed first order kinetics and depended upon protein concentration. BML was more radiosensitive than HML and EWL. Comparisons of the ultraviolet scans of the native and irradiated enzymes indicated that tryptophan was being destroyed in HML as it is known to occur in EWL. Hydroxy radical scavengers, cysteine and t-butanol, afforded radioprotective action for the milk lysozymes. In a dry state, both the milk lysozymes were less sensitive to -irradiation than in aqueous solution, requiring Mrad dosages for partial inactivation compared to Krad dosages for the same extent of inactivation in aqueous solution. However, as was observed in aqueous solution and also in the dry state, BML was more radiosensitive than HML and EWL.


ISOLATION AND PURIFICATION OF Clostridium perfringens ENTEROTOXIN BY AFFINITY CHROMATOGRAPHY

June 2008

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44 Reads

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3 Citations

Sporulating cells of certain strains of Clostridium perfringens produce an intracellular enterotoxin believed to be the causative agent of C. perfringens food poisoning. Rabbit antibody preparations to both vegetative and sporulating cell extracts were used to purify toxin by differential immuno-affinity chromatography. Solid phase immune adsorbents were prepared by attaching antibody via α-amino groups to succinylaminoethyl Sepharose-4B to give two resins: one binding anti-vegetative immunoglobulin G (IgG), (V-resin), the other, antisporulating IgG, (S-resin). Sporulating cell extract was passed through resin with bound IgG to vegetative cell extract. A large protein fraction representing antigens common to both C. perfringens forms was retained, but toxin was not measurably absorbed to the resin as determined by erythemal activity in rabbits. The toxic fraction was then passed through resin with bound IgG to sporulating cell extract. The majority of the protein did not adhere to this resin; however, that which did, showed erythemal activity in rabbits. Disc-gel electrophoresis of the protein fraction eluted from resin with bound IgG to sporulating cell extract demonstrated the presence of five components. One elicited erythemal activity in rabbits. The maximum capacity of the V-resin column was 102 μg of bound protein/ml of resin, and of the S-resin column, 32 μg of bound protein/ml of resin. A 150-fold purification was achieved by the procedure. Resins could be used repeatedly.


Effects of Processing and Storage on the Water‐Soluble Vitamin Content of Human Milk

August 2006

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308 Reads

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17 Citations

Effects of four heat treatments (62.5°C, 30 min; 72°C, 15 sec; 88°C, 5 sec; and 100°C, 5 min) and frozen storage (−20°C, 4 wk) on water-soluble vitamin content of composite samples of mature human milk were determined. No changes in riboflavin, biotin, and total pantothenic acid content were observed after any of the treatments. Heating at 100°C for 5 min decreased thiamin, vitamin B12, and vitamin C contents. Vitamin Bs content was reduced by heating at 88°C and 100°C. Free folate was not affected by heating while total folate concentration was decreased similarly by all heat treatments. During frozen storage at −20°C, niacin and free pantothenic acid concentrations were reduced; riboflavin, vitamin Bs, biotin, total pantothenic acid, vitamin C, and vitamin B12 were unaffected; while measurable thiamin and free folate concentrations increased. Overall, the two lower temperature treatments were less detrimental to the water-soluble vitamins in human milk.


Low dose gamma irradiation of bovine metmyoglobin

August 2006

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11 Reads

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37 Citations

SUMMARY Bovine metmyoglobin (MetMb) in a pure state, in several differing states of purity and in meat was irradiated with ionizing gamma rays. Irradiation caused the brown MetMb to convert to a red myoglobin pigment which is similar but not identical to oxymyoglobin (MbO2). The formation of red pigment was accelerated at slightly basic pHs and under nitrogen atmospheres. Rate of red pigment formation is not dependent upon MetMb purity. Rate of red pigment formation in meat is identical to the rate of formation in buffered MetMb solutions. This study also demonstrated that gamma irradiation does alter MetMb structure. This alteration may be the loss of amide nitrogen from the protein.


Thermal Destruction of Immunoglobulin A, Lactoferrin, Thiamin and Folic Acid in Human Milk

August 2006

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61 Reads

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14 Citations

J. N. MORGAN

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R. T. TOLEDO

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R. R. EITENMILLER

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[...]

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F. MADDOX

Kinetic parameters for thermal destruction of immunoglobulin A (IgA), lactoferrin, thiamin and folic acid in human milk were determined. Degradation proceeded following first order reaction kinetics. The times for 90% degradation (D value) at 60°C were, in seconds, 4.9 × 104 (68°C C 78°C) for IgA; 2.4 × 103 (58°C 70°C) for lactoferrin; 7.7 × 105 (95°C C 110°C) for thiamin and 1.9 × 104 (62°C C 78°C) for folic acid based on inactivation data at four constant temperatures between the range indicated. Z values (temperature change to alter degradation rate by a factor of 10) were 5.5°C, 4.7°C, 28.4°C, and 6.4°C for IgA, lactoferrin, thiamin and folic acid, respectively.


Use of gamma irradiation to prevent aflatoxin in bread

August 2006

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17 Reads

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11 Citations

Irradiation doses of 100 and 200 Krad reduced the growth of Aspergillus parasiticus in bread stored for 10 days at 254DEC. With bread stored longer than 2 wk, mold growth in irradiated samples tended to approach the amount which occurred in the non-irradiated controls. Mold strain NRRL 3000 seemed slightly more sensitive to irradiation-than strain NRRL 2999. A 200 Krad dose prevented anatoxin production by either strain at both high and low levels of inoculation with spores in bread stored for 10 days, except in one case where a very low level of anatoxins was detected. At 100 Krad, bread inoculated with 102 spores/ slice contained none to low concentrations of anatoxins after storage for 10 days, but with 106 spores/slice higher amounts of toxins were detected. When ihe inoculated bread was stored for periods up to 6 wk, the 200 Krad treatment prevented aflatoxin production in all bread samples containing 10/sup 2/ spores/ slice, and in almost all samples containing 10/sup 6/ spores/ slice. The 100 Krad treatment also prevented aflatoxin production during 6 wk of storage in bread that contained 10/sup 2/ spores/ slice. However, with 10/sup 6/ spores/ slice both strains were capable of producing very highmore » amounts of aflatoxins after irradiation nt 100 Krad. At 1, 2 and 6 wk of storage, these amounts were greater than the unirradiated controls, suggesting possible stimulation of aflatoxin production. (auth)« less


Fig. 1. Immunofluorescent detection of Campylobacter in poultry flock house watering systems. Springloaded stainless steel pins taken from the drinking nipples were examined for the presence of Campylobacter in the biofilm with fluorescently labeled antibody. Negative samples exhibited little background fluorescence (A). Samples containing Campylobacter cells exhibited areas of bright fluorescence (B). Campylobacter cells were detected in the nipple drinkers during times when the flock was culture positive. Campylobacter was not detected in the waterers during periods when birds were culture negative.
Fig. 2. RAPD types resulting from amplification of DNA template from Campylobacter jejuni isolates from three sequential broiler flocks and their drinking water. (A) Whole cell template was amplified with the primer OPA 11 and amplicons were separated on a 1.5% agarose gel containing ethidium bromide. RAPD types II and III were associated with strains cultured only from the nipple waterers whereas types IV, V, VI, and VII were associated with strains cultured from the chickens. (B) Because type I was found in isolates cultured from both chickens and  
Fig. 1. Immunofluorescent detection of Campylobacter in poultry flock house watering systems. Springloaded stainless steel pins taken from the drinking nipples were examined for the presence of Campylobacter in the biofilm with fluorescently labeled antibody. Negative samples exhibited little background fluorescence (A). Samples containing Campylobacter cells exhibited areas of bright fluorescence (B). Campylobacter cells were detected in the nipple drinkers during times when the flock was culture positive. Campylobacter was not detected in the waterers during periods when birds were culture negative. our sample size, and the C. jejuni strain may have been present in drinking water during the growth of flocks 2 and 3 but was culturable only during the growth of flock 1. Campylobacter can exist in a viable but nonculturable  
Detection of Campylobacter jejuni Strains in the Water Lines of a Commercial Broiler House and Their Relationship to the Strains That Colonized the Chickens
  • Article
  • Full-text available

January 2003

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114 Reads

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96 Citations

Avian Diseases

Campylobacter jejuni is frequently present in the intestinal tract of commercial broiler chickens, and their drinking water has been proposed to be an initial source of bacteria for newly hatched chicks. We studied three sequential commercial broiler flocks raised in a house from which we had cultured C. jejuni from the nipple waters prior to placement of the first flock. Campylobacter cells were detected by immunofluorescence in the biofilm of the drinking nipples during the weeks when the flock was colonized with C. jejuni but not during weeks when the birds were negative. Campylobacter jejuni was isolated from the drinking water during the growth of the first flock and was present in the birds from all three flocks. Randomly amplified polymorphic DNA (RAPD)-polymerase chain reaction (PCR) typing with primer OPA11 indicated that seven distinct strains were present within the broiler house. One strain found in drinking water was similar to a strain found in birds in the second flock; however, RAPD-PCR with primer HLW85 showed that the strains were not identical. These results suggest that although the watering system is a potential source of C. jejuni in broiler flocks, the waterborne strain in this study was not detected in the birds.

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FIG. 1. Prevalence of C. jejuni recovered from cecal droppings in flocks 1 ( n ϭ 67) and 2 ( n ϭ 28). 
FIG. 2. RAPD-PCR profiles of C. jejuni . 
FIG. 3. 23S rRNA-PCR of C. jejuni . 
Molecular Epidemiology of Campylobacter jejuni in Broiler Flocks Using Randomly Amplified Polymorphic DNA-PCR and 23S rRNA-PCR and Role of Litter in Its Transmission

February 1999

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78 Reads

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51 Citations

Poultry has long been cited as a reservoir for Campylobacter spp., and litter has been implicated as a vehicle in their transmission. Chicks were raised on litter removed from a broiler house positive for Campylobacter jejuni. Litter was removed from the house on days 0, 3, and 9 after birds were removed for slaughter. Chicks were raised on these three litters under controlled conditions in flocks of 25. None of these birds yielded C. jejuni in their cecal droppings through 7 weeks. Two successive flocks from the same Campylobacter-positive broiler house were monitored for Campylobacter colonization. Campylobacter jejuni prevalence rates were determined for each flock. Randomly amplified polymorphic DNA (RAPD)-PCR and 23S rRNA-PCR typing methods were used to group isolates. A high prevalence (60%) of C. jejuni in flock 1 coincided with the presence of an RAPD profile not appearing in flock 2, which had a lower rate of prevalence (28%). A 23S rRNA-PCR typing method was used to determine if strains with different RAPD profiles and different prevalence rates contained different 23S sequences. RAPD profiles detected with higher prevalence rates contained a spacer in the 23S rRNA region 100% of the time, while RAPD profiles found with lower prevalence rates contained an intervening sequence less than 2% of the time. Data suggest varying colonizing potentials of different RAPD profiles and a source other than previously used litter as a means of transmission of C. jejuni. These molecular typing methods demonstrate their usefulness, when used together, in this epidemiologic investigation.


Accumulation of Elements in Fractionated Broiler Litter during Re-Utilization

March 1998

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12 Reads

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9 Citations

The Journal of Applied Poultry Research

Poultry litter is a mixture of broiler (chicken, Gallus gallus domesticus) excreta and bedding material. The broiler industry may produce more litter than can be locally land-applied without causing environmental degradation. Passing whole litter through 0.83- and 3.33-mm sieves separates a fine fraction that can be land-applied as fertilizer and a coarse fraction that can be mixed with bedding and re-utilized in broiler houses. A study was conducted to determine whether the coarse litter fraction could be re-utilized as a supplement for bedding in broiler houses without significant accumulation of heavy metals and other elements. Wood shavings supplemented with either stored or fresh coarse litter and a treatment of unsupplemented wood shavings were used as bedding material during a 40-day broiler growth period. Initial and final litter samples were collected and analyzed for Al, As, B, Ba, Ca, Cd, Co, Cr, Cu, Fe, K, Mg, Mn, N, Na, Ni, P, Pb, Si, Sr, and Zn. Results indicated that element concentrations in wood shavings bedding supplemented with litter did not increase significantly during the final 30 days of a 40-day growth period. However, significant accumulation of several litter elements in unsupplemented wood shavings bedding during the entire 40-day growth period indicated that repeated recycling of litter as a bedding supplement may lead to significant accumulation of elements in litter.


Antibiotic Resistance of Bacterial Litter Isolates

March 1998

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45 Reads

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100 Citations

Poultry Science

Use of antibiotics in subtherapeutic doses as growth-promoting feed additives for animal production is widespread in the U.S. and throughout the world. Previous studies by our research group concluded that size fractionation of poultry (broiler) litter followed by storage facilitated reutilization of litter as a soil amendment or bedding supplement. However, litter microbial contamination, including antibiotic-resistant populations, and accumulation of metals and other elements may limit litter reutilization. Litter from four broiler houses was separated into a fine fraction for use as a soil amendment, and a coarse fraction for reutilization as a bedding supplement in growing subsequent flocks of broilers. Fractions and whole litter were stored in indoor piles simulating farm storage conditions for 4 mo with periodic analysis for metals, other elements, and culturable bacteria (including total and fecal coliform, Aeromonas hydrophila, Pseudomonas aeruginosa, Yersinia enterocolitica, and Campylobacter jejuni). Representative bacterial isolates were tested for their sensitivity to 12 common antibiotics (ampicillin, bacitracin, cephalothin, erythromycin, gentamicin, kanamycin, nalidixic acid, neomycin, penicillin, streptomycin, sulfisoxazole, and tetracycline) using the Kirby-Bauer technique. Pathogens and indicator bacteria tested were found to be resistant to multiple antibiotics. Data suggest that microbial contamination of litter should be reduced or eliminated prior to reutilization to minimize environmental health risks related to transfer of antibiotic-resistant bacteria to humans or other animals.


Citations (36)


... Calcium is a critical mineral for infant development, particularly for bone growth and mineralization, as well as for muscle function and nerve signaling [65][66][67][68][69]83]. The variability in calcium concentrations observed in this study likely reflects individual differences in maternal diet, calcium supplementation, and the stage of lactation, all of which can influence the amount of calcium present in breast milk. ...

Reference:

Determination of Minor and Trace Elements in Breast Milk of Lactating Mothers in Early Lactation from Tehran, Iran Using Neutron Activation Analysis Method
Calcium, Phosphorus, and Magnesium Contents of Human Milk During Early Lactation
  • Citing Article
  • May 1983

Journal of Pediatric Gastroenterology and Nutrition

... A total of 6 Genera were isolated from the Laxmibazar Overhead tank. Among them, 4 were Grampositive (S. aureus, Bacillus, Micrococcus, Enterococcus), and 2 were Gram-negative bacteria (Morgan, 1988). ...

Thermal Destruction of Escherichia coli and Klebsiella pneumoniae in Human Milk

Journal of Food Protection

... In Japan, S. Infantis was isolated from both eggshells and egg contents [47]. The isolation of S. Infantis from the ovaries of commercial laying hens during slaughter [48] suggested that the organism could be transmitted to the egg through transovarian infection and also caused public health concern [49]. However, there is currently insufficient data on this subject. ...

Prevalence of Salmonella enteritidis and Other Serovars in Ovaries of Layer Hens at Time of Slaughter

Journal of Food Protection

... Antibiotic doses used were as follows: kanamycin (30 µg), tetracycline (30 µg), erythromycin (15 µg), ampicillin (25 µg), tobramycin (10 µg), streptomycin (10 µg), rifampicin (5 µg), and chloramphenicol (30 µg). According to Barnhart and Pancorbo (1992), the antibiotic doses were chosen. Antibiotic resistances were determined by comparing bacterial isolate inhibition zone diameters with NCCLS criteria (NCCLS, 2004). ...

Cytotoxicity and Antibiotic Resistance Profiles of Aeromonas hydrophila Isolates From a Broiler Processing Operation

Journal of Food Protection

... It is made immunologically relevant antigens before the infant gains the ability to generate their own IgA response, reducing the risk of enteric infection and necrotizing enterocolitis [15][16][17][18] When the maternal milk containing these valuable factors is unavailable, Donor human milk and human milk-derived fortifiers are increasingly utilized for VLBW infants rather than formula [19,20]. Donor milk can be expressed throughout a donor's lactation cycle, often months following parturition when the concentrations of many milk factors, including EGF and IgA, are lower than immediately after birth [21][22][23][24]. We have observed in a preclinical model that milk expressed closer to parturition contained more EGF and offered more protection from enteric pathogens than milk expressed later in lactation [14]. ...

IgA, IgG, IgM and Lactoferrin Contents of Human Milk During Early Lactation and the Effect of Processing and Storage

Journal of Food Protection

... Traditionally, bacteria including coliforms and enterococci have been used to estimate the level of faecal contamination of water and/or shellfish (Oliveira et al., 2011;Pancorbo and Barnhart, 1992), and may be referred to collectively as Faecal Indicator Bacteria (FIB). In Europe, Escherichia coli is adopted as the traditional indicator of faecal (sewage) contamination in shellfish and is used for risk assessment and management purposes (EC, 2004). ...

Microbial pathogens and indicators in estuarine environments and shellfish: Critical need for better indicator(s) of human-specific fecal pollution
  • Citing Article
  • January 1992

... Significantly decreased Aeromonas hydrophila, but refrigeration at 1.1 C for 48 hours significantly increased it. 28 Oxidizing-When A. hydrophila was exposed to oxidising raw ground fluids with high Fe2+ levels (460-1.070 lmol), it immediately dropped by 2-3 log unit. ...

Recovery of Aeromonas hydrophila from Carcasses and Processing Water in a Broiler Processing Operation

Journal of Food Protection

... Understanding broiler management factors that could be used to predict the occurrence of food-borne pathogens in reused litter is vital to limiting their prevalence in post-harvest chicken production. Although there are studies on the microbiology of reused litter, these studies have been limited to wood shavings and rice-hull based litter 1 (Kelley et al., 1995;Brooks et al., 2010;Roll et al., 2011;Volkova et al., 2011;Chinivasagam et al., 2012;Eberle et al., 2013;Roberts et al., 2013;Wei et al., 2013;Brooks et al., 2016;Rauber Wurfel et al., 2019;Voss-Rech et al., 2019;Valeris-Chacin et al., 2021;Chinivasagam et al., 2022). Roberts et al. (2013) demonstrated that commensals (Staphylococcus spp., Enterococcus spp., and Clostridium perfringens) and pathogens (Salmonella and Listeria monocytogenes) were spatially and temporally distributed in 8 broiler houses. ...

Bacterial Pathogens and Indicators in Poultry Litter during Re-Utilization

The Journal of Applied Poultry Research