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Detection of Salmonella spp. in eggs: DNA analyses, culture techniques, and serology.

A polymerase chain reaction (PCR) method for direct detection of Salmonella spp. in whole-shell eggs is described. The method does not require isolating strains. Preenrichment, rapid DNA isolation, and a nested PCR system targeting the invA gene enabled detection of the genus Salmonella. The specific nested PCR product of 283 base pairs was formed from all 21 serovars, including 43 Salmonella strains tested. No PCR product was formed from 56 non-Salmonella enterobacteriacea and other bacterial strains tested. Experiments with artificially contaminated eggs showed a detection limit of about 10(3)-10(4) colony-forming units (cfu)/egg before and about 1-10 cfu/egg after preenrichment. In analysis of 180 single eggs from 4 flocks and 36 pools of 5 eggs each from another 4 flocks from the same producer, Salmonella spp. were detected in 5 of 90 eggs from 2 different flocks. Determination of anti-Salmonella antibodies in eggs yielded positive results for 2 additional and the 2 PCR-positive flocks. In contrast, classical selective culture detected Salmonella spp. in only 1 of 100 eggs in one flock when 100 eggs from each flock were analyzed.

Bacteriological Techniques↗

Outbreaks of egg drop syndrome due to EDS-76 virus in quail (Coturnix coturnix japonica).

Two outbreaks of the egg drop syndrome were observed in quail flocks maintained on a farm together with chickens. The decrease in egg production ranged from 10.6 per cent to 50.6 per cent, and the number of soft-shelled eggs increased with the decline in egg production. In both the outbreaks haemagglutination inhibiting antibodies to EDS-76 virus were detected. Fluorescent viral antigen specific to EDS-76 virus was also detected in the lining epithelial and glandular cells of the uterus.

Adenoviridae Infections↗

Capillaria procyonis (Nematoda: Trichuroidea) eggs from the tongue of the raccoon (Procyon lotor).

During an investigation to determine the prevalence and intensity of larvae of Trichinella spiralis in raccoons (Procyon lotor), unembryonated capillariid-type eggs were found routinely in the sediment resulting from 1% pepsin-HCl acid digests of the tongue. The sediment from 36 (90%) of the 40 tongue digests examined contained these capillariid-type eggs. Mean measurements of egg width and length were 25 +/- 2 microns and 62 +/- 2 microns, respectively. In addition, the surface of the egg shell had a pitted appearance. Adult parasites were not found in any of the tongue digests. The egg measurements and surface topography agree with what has been described previously and are tentatively identified as Capillaria procyonis. The detection of C. procyonis eggs in the present report indicates that this parasite inhabits the epithelial lining of the raccoon tongue, in addition to the esophagus, and that eggs expelled by females are deposited in the tracts these parasites create as they migrate.

Animals↗

Developmental and age-related changes in proteins in the female reproductive tract of Heligmosomoides polygyrus (Nematoda).

Proteins in the female reproductive tract of Heligmosomoides polygyrus at days 8, 16, 35, 90, and 140 postinfection (PI) were examined using polyacrylamide gel electrophoresis. Sixteen-day-old and 140-day-old worms also were examined histochemically. Egg production of these worms was assessed for each age group. In analyzing proteins using electrophoresis, the reproductive tracts were separated into 3 sections: the tip, or anteriormost part of tract, containing oogonia; the middle region, containing developing oocytes; and the posterior region, containing the uterus with fertilized eggs. Three major reproductive tract proteins were identified as having molecular weights of greater than 140 kDa, 115 kDa, and 82 kDa. These were found in all parts of the reproductive tract from worms of all ages except those at 8 days PI (which are too young to produce eggs) and are believed to be yolk proteins. Four low molecular weight proteins (L1-4) are believed to be nucleoproteins. L4 was absent from the posterior section of the reproductive tracts and L3 was limited to the posterior sections and may be associated with sperm stored in the uterus. Of 5 high molecular weight proteins the second heaviest, designated H2, appeared to be relatively more concentrated in the posterior sections of the reproductive tract. An 85-kDa protein was limited to the tip and middle sections of reproductive tracts. Histochemical tests on sectioned H. polygyrus showed strong positive reactions for protein in cytoplasmic granules in developing oocytes and in eggs of younger worms (16 days) but a reduced reaction in older worms (140 days). Strains for collagen showed a slight positive reaction in and between developing oocytes and a strong reaction in the egg shells. Stains for nucleoproteins particularly reacted with sperm stored in the uterus, and slightly reacted with fertilized eggs and the nucleoli of the intestinal and ovarian epithelium. Egg production by H. polygyrus increased to 123 eggs/female/day by 16 days PI but declined from 121 eggs/female/day at 35 days PI to 64 eggs/female/day in worms 140 days old. Electrophoresis indicated no loss in the different types of proteins in the reproductive tract of older worms, but histochemistry and protein content assays suggest that older worms that produce fewer eggs contain a relatively smaller amount of protein in the female reproductive tract.

Aging↗

Efficacy of electrolyzed oxidizing water for the microbial safety and quality of eggs.

During commercial processing, eggs are washed in an alkaline detergent and then rinsed with chlorine to reduce dirt, debris, and microorganism levels. The alkaline and acidic fractions of electrolyzed oxidizing (EO) water have the ability to fit into the 2-step commercial egg washing process easily if proven to be effective. Therefore, the efficacy of EO water to decontaminate Salmonella Enteritidis and Escherichia coli K12 on artificially inoculated shell eggs was investigated. For the in vitro study, eggs were soaked in alkaline EO water followed by soaking in acidic EO water at various temperatures and times. Treated eggs showed a reduction in population between > or = 0.6 to > or =2.6 log10 cfu/g of shell for S. Enteritidis and > or =0.9 and > or =2.6 log10 for E. coli K12. Log10 reductions of 1.7 and 2.0 for S. Enteritidis and E. coli K12, respectively, were observed for typical commercial detergent-sanitizer treatments, whereas log10 reductions of > or =2.1 and > or =2.3 for S. Enteritidis and E. coli K12, respectively, were achieved using the EO water treatment. For the pilot-scale study, both fractions of EO water were compared with the detergent-sanitizer treatment using E. coli K12. Log10 reductions of > or = 2.98 and > or = 2.91 were found using the EO water treatment and the detergent-sanitizer treatment, respectively. The effects of 2 treatments on egg quality were investigated. EO water and the detergent-sanitizer treatments did not significantly affect albumen height or eggshell strength; however, there were significant affects on cuticle presence. These results indicate that EO water has the potential to be used as a sanitizing agent for the egg washing process.

Animals↗

Salmonella enteritidis phage type 4 infection: association with hen's eggs.

Epidemiological and microbiological investigation of the four outbreaks of Salmonella enteritidis phage type (PT) 4 that occurred in Wales in 1988 implicated foods containing shell eggs from hens as the vehicle of infection. In a case-control study of sporadic cases there was a significant association between infection and egg consumption in the 3 days before illness. These findings provide data to support the warnings from the Departments of Health in the United Kingdom about risks of infection from eating raw or undercooked hens' eggs.

Adult↗

Genetic analysis of two female-sterile loci affecting eggshell integrity and embryonic pattern formation in Drosophila melanogaster.

We have analyzed female-sterile mutations at the X-linked loci fs(1)Nas and fs(1)ph which show allele-specific effects on egg shell structure and embryonic pattern formation. The majority of mutant alleles at both loci lead to a collapsed egg phenotype. The maternal effect lethal phenotype is characterized by cuticle defects resembling those found in three autosomal mutants of the terminal class. We have analyzed the complementation behavior of various heteroallelic combinations at both loci and show that one such combination at the fs(1)Nas locus is capable of restoring normal fertility. We have investigated possible interactions between fs(1)Nas and fs(1)ph and also between the terminal allele of fs(1)Nas and various maternal effect mutations altering the anteroposterior polarity of embryos. We have isolated one new allele of fs(1)Nas which combines the locus-typical phenotypic features with novel cuticle phenotypes. Our results suggest that the products of fs(1)Nas and fs(1)ph are required for the stability of the vitelline membrane and are also involved in a morphogenetic pathway necessary for the correct differentiation of the terminal regions of the embryo. Possible mechanisms to account for the association of these two functions are discussed.

Alleles↗

The in vivo development of Schistosoma bovis in the Nile rat.

The in vivo development of the Sudanese strain of Schistosoma bovis in the Nile rat, Arvicanthus niloticus is described. Six stages in development are distinguished: the 'lung form' (stage 1), attaining maximum numbers on day 5 post-infection; the 'closed-gut form' (stage 2) on day 14, characterized by the union of the gut caeca behind the ventral sucker; 'organogeny' (stage 3) on day 17, the male possessing one testis and a gynaecophoric canal and the female a narrow uterus; 'gametogeny' (stage 4) on day 26, with pairing, the male having four fully developed testes and the female an ovary; 'egg-shell formation' (stage 5) on day 35; 'oviposition' (stage 6) on day 37, with the female showing uterine eggs. The time to complete in vivo development in the Sudanese strain of S. bovis is more rapid than in the Israeli strain, 37 and 42 days respectively.

Animals↗

Biology of Schistosoma japonicum. From cercaria penetrating into host skin to producing egg.

The biology of the Chinese mainland strain of Schistosoma japonicum in mammalian phase of the life cycle has been studied in the past three decades. The mode of cercarial penetration is found to involve enzyme secretion and movement. The entry of schistosomulum into the blood vessel in the hypodermal connective tissue is evidenced after skin penetration. The growth and development of this fluke in mice and rabbits are investigated by physiological, histochemical and ultrastructural methods. The development of the fluke may be divided into eight stages. In the definitive host-range, at least 45 species and subspecies of 34 genera in 18 families of 7 mammalian orders have been demonstrated, and the susceptibility to infection in domestic animals is compared. The process of egg formation and embryonic development of this fluke are studied histologically and histochemically. It is suggested that the chemical nature of the egg shell should be a quinontanned protein system. The characteristics and dynamic changes of cytochemical substances in embryonation of eggs are described.

Animals↗

The effects of ingested petroleum on oviposition and some aspects of reproduction in experimental colonies of mallard ducks (Anas platyrhynchos).

Compared to unmated mallard ducks fed an uncontaminated diet, unmated birds given food contaminated with 3 ml South Louisiana crude oil per 100 g dry weight showed an 84% decline in the daily rate of oviposition, a 33% decrease in egg-shell thickness and at autopsy more than 82% of the ovarian mass consisted of atretic follicles. Similar studies on groups of mated females showed that although the addition of 1 ml South Louisiana crude oil/100 g dry food had no effect on the daily rate of oviposition, none of the eggs had been fertilized while a concentration of 3 ml South Louisiana crude oil/100 g dry food suppressed the daily rate of oviposition significantly. Less than 25% of these eggs had been fertilized and only 40% of the fertilized eggs yielded viable ducklings. In both of these groups of mated birds, normal patterns of oviposition, fertilization and hatchability were restored after removal of petroleum from the diet. The addition of 1 ml Kuwait crude oil/100 g dry food had no effect on the rate of oviposition, the incidence of fertility or the hatchability of the fertilized eggs. The addition of 3 ml oil/100 g dry food completely abolished oviposition, but a normal rate of oviposition was restored when the concentration of the crude oil was reduced from 3 to 1 ml/100 g dry food. However, the incidence of fertilization remained low and none of the fertilized eggs gave rise to viable ducklings. Kuwait crude oil had no effect on shell thickness.

Animals↗

Potential of alfalfa as an alternative molt induction diet for laying hens: egg quality and consumer acceptability.

Dietary molt induction to initiate additional egg laying cycles in commercial laying hen flocks is a wide spread practice in the United States. Feed deprivation is the most commonly used method but this practice has generated several concerns which has lead to research for viable alternative approaches. From a management standpoint a single ingredient molting diet consisting of high fiber-low energy represents an easily adaptable diet for large laying hen production units. Alfalfa meal is readily available in most commercial locations and possesses many of the desirable properties of an ideal laying hen molt diet. In the current study hens at a commercial laying facility were molted by both alfalfa and feed deprivation. After the hens had reentered post-molt commercial egg production, eggs were examined for egg quality performance. Egg shell strength, albumen height, yolk height, weight, length, and yolk color were all tested using various mechanical techniques. The eggs were also sampled for testing by consumer sensory panels that assessed the desirability of the eggs' color and flavor/texture. Eggs laid by hens molted by alfalfa had a significantly lower (p<0.05) "a*" level of colorimetry. Eggs laid by hens molted with alfalfa also exhibited significantly higher (p<0.05) egg weights and length. In the consumer sensory test, there was no significant difference (p>0.05) in color or flavor/texture scores in eggs from either feed deprived or alfalfa molted hens. The consumer sensory and mechanical quality attributes indicates that alfalfa shows promise as an alternative molt induction diet by providing a single diet option for extending egg production into a second egg laying cycle.

Animal Feed↗

Reproductive performance of hens fed field beans and potential relationships to vicine metabolism.

Field beans (faba beans, broad beans, horse beans, ackerbohnen, Vicia faba L.) depress weight of eggs when fed to laying hens. Vicine isolated from field beans has been shown to depress egg weight and hatchability when fed in large amounts. The present study was undertaken to determine the effect of using three varieties of field beans, varying in tannin and total vicines content, at 20% in breeder diets on reproductive performance of hens and associated vicine transfer from the diet to the egg. Data on egg production, feed consumption, egg weight, body weight, egg quality, hatchability, and hen blood chemistry were obtained. Direct spectrophotometric and high pressure liquid chromatographic (HPLC) analyses of field beans, egg albumen, and egg yolk were conducted. Performance of the hens was excellent on all production characteristics except that there was a nonsignificant reduction in egg weight, a significant reduction in egg shell thickness for hens fed all three types of field beans, and a highly significant reduction in hatchability of eggs from hens fed one but not the other two sources of field beans. The hatchability effect was not correlated with vicine or tannin content of the field beans and only traces of vicine were transferred to the egg by the hen from the diet. Although larger amounts of vicine depressed hatchability when injected into fertile eggs, the trace amounts found in eggs from feeding field beans did not account for the reduction in hatchability observed in this study. It is possible that the hydrolysis product of vicine, divicine or the analogous product from convicine, isouramil, or both, may be involved in toxicity, because these substances are known to be the cause of a hemolytic anemia in some humans that suffer from favism due to consumption of field beans.

Animal Feed↗

Embryos and embryonic envelopes in eggs of two cyclophyllidean cestodes.

Membranes and envelopes around the embryos in eggs of Cotugnia digonopora and Raillietina (R.) echinobothrida have been studied in detail. This is the first study giving details of the egg-shell of a Cotugnia species. There are 3 membranes and an equal number of envelopes around the embryos in eggs of C. digonopora , while there are 2 membranes and only one envelope in case of eggs of R. echinobothrida . Noteworthy is the presence of a membrane in eggs of C. digonopora , termed "middle capsule", between and separating the 2 envelopes viz. the outer and inner. The membranes and envelope around the embryos in eggs of R. echinobothrida have been found to be identical with those of R. galeritae .--The 3 pairs of embryonic hooks of the 2 cyclophyllideans studied are collared --a feature of specific importance.

Animals↗

Growth of Salmonella enteritidis in artificially contaminated eggs: the effects of inoculum size and suspending media.

Growth profiles of two isolates of Salmonella enteritidis phage type (PT) 4 inoculated into either the albumen of whole shell eggs or into separated albumen were found to be markedly affected by the size of the inoculum and the composition of the medium used to suspend the cells prior to inoculation. Using our model with an inoculum of two cells, multiplication of the Salmonella was not seen in 93% of eggs held at 20 degrees C for 8 days. In approximately 7% of eggs, however, growth occurred during the 8 days of storage. If the inoculum equaled or exceeded 25 cells per egg when eggs were subsequently stored at 20 degrees C, or 250 cells per egg when eggs were stored at 30 degrees C, high levels of growth of Salmonella in the egg occurred significantly more frequently than when the inoculum was two cells. High levels of growth were also seen more frequently if the inoculum was suspended in buffered peptone water or maximal recovery diluent rather than in phosphate buffered saline. Growth of Salmonella in separated albumen occurred very infrequently (1.1% of samples) at low inoculum levels and did not become significant until the inoculum was 250 cells or greater. Growth in the albumen was unaffected by the composition of the suspending medium. Provided that the inoculum was approximately 2 cells per egg and the bacteria were suspended in PBS, observed growth profiles of S. enteritidis inoculated into the albumen of whole eggs resembled those in naturally contaminated eggs.

Animals↗

Self-selection of calcium during the rearing and early laying periods of White Leghorn pullets.

The ability of pullets to selectively consume calcium to meet the requirements of the rearing and early laying period was studied in White Leghorns. In the first of two experiments, 456 pullets were fed either an .89% calcium rearing diet (control) or a .35% calcium diet in combination with a separate source of 38% calcium chick-sized grit (calcium self-selection) from 35 to 126 days of age. Twenty-four pullets from each of these rearing treatments were selected for the second experiment at 112 days of age and were housed in individual laying cages. Rearing treatments were continued until 176 days of age with mash and grit consumption recorded daily for each pullet. From 177 to 225 days of age these pullets were fed a 3.50% calcium diet containing either ground limestone or oyster shell as a calcium source or a .35% calcium diet fed in combination with a separate source of oyster shell. Pullet calcium intake reflected the requirements for growth during the rearing period, calcium storage in the medullary bone 19 days prior to lay, and egg shell formation during the laying period; average calcium intake for these periods was 1.18, 2.08, and greater than 3.50%, respectively. Pullets also demonstrated the ability to compensate for rearing deficiencies by increased calcium consumption when fed oyster shell on an ad lib basis. Calcium consumption was found to be significantly higher on days when an oviposition occurred than when no eggs were laid. Mash consumption during the early laying period also reflected the reproductive status of the hen with the highest consumption on days when both oviposition and ovulation occurred, intermediate consumption when either oviposition or ovulation occurred, and lowest consumption when neither oviposition nor ovulation was detected.

Aging↗

Two new Huffmanela species, H. japonica n. sp. and H. shikokuensis n. sp. (Nematoda: Trichosomoididae), from marine fishes in Japan.

Two new species of trichuroid nematodes. Huffmanela japonica n. sp. and Huffmanela shikokuensis n. sp., are established on the basis of their egg morphology and biological characters; the eggs of both species occur in the musculature of marine fishes from the Inland Sea of Japan. The dark-shelled eggs of H. japonica are found locally in masses ("black spots") in the flesh of Upeneus bensasi (Temminck et Schlegel) (Mullidae, Perciformes) and are characterized mainly by their shape and size (58-69 x 26-30 microm), an aspinose superficial transparent envelope enclosing the egg proper, relatively small polar plugs, and by their thick egg wall (4-5 microm). The eggs of H. shikokuensis are also dark-colored and are found evenly distributed in the musculature of Stephanolepis cirrhifer (Temminck et Schlegel) (Monacanthidae, Tetraodontiformes); they are characterized mainly by their shape and size (78-90 X 36-45 microm), by a very thin and aspinose superficial transparent envelope, large polar plugs, and relatively thin egg wall (3 microm). Histological sections of the host's infected musculature showed the presence of H. shikokuensis nematodes inside the muscle cells and in the intercellular spaces. A key to Huffmanela species based on egg morphology is provided.

Animals↗

In vitro study of Salmonella enteritidis and Salmonella typhimurium definitive type 104: survival in egg albumen and penetration through the vitelline membrane.

Salmonella enteritidis and Salmonella typhimurium definitive type 104 (DT104) have been detected in the chicken oviduct, and their survival in egg albumen at the chicken body temperature of 42 degrees C may be important in oviductal and transovarian contamination of intact shell eggs. Eight S. enteritidis and 24 S. typhimurium DT104 strains were tested for their in vitro survival in egg albumen. The concentration of the organisms declined more rapidly when incubated at 42 degrees C than at 37 degrees C and dropped to nondetectable levels within 96 h at the higher, but not at the lower, temperature. In another experiment, 3 S. enteritidis and 3 S. typhimurium DT104 strains were randomly selected, and dosages of 20 and 200 cells of each strain were inoculated onto the vitelline membranes of egg yolks, which were then submerged in the original albumen and incubated for 24 h at 42 degrees C. Under these conditions, the organisms survived in albumen but did not penetrate the vitelline membrane. However, in a similar experiment, penetration did occur when the specimens were incubated at 30 degrees C for 72 h. The results suggest that low numbers of S. enteritidis and S. typhimurium DT104 can be expected to survive in egg albumen during the 24-h period of egg formation in the oviduct but would be unlikely to invade the yolk.before oviposition. However, depending on storage conditions following oviposition, S. enteritidis, as well as S. typhimurium DT104, could survive longer and may eventually invade the egg yolks.

Albumins↗

Thermostabilization of ovalbumin in a developing egg by an alkalinity-regulated, two-step process.

Native ovalbumin has been known to convert into a heat-stable form, S-ovalbumin, either in an avian shell egg or in an isolated ovalbumin solution. Recently, similar conversion of ovalbumin in fertile eggs was also reported. We found that the conversion into S-ovalbumin was slower in fertile eggs than in unfertile eggs under the same incubation conditions on the basis of calorimetric analyses for the samples isolated from those eggs. During the incubation, there were differential pH changes of white in the fertile and unfertile eggs. When the pH of purified ovalbumin was manually adjusted so as to simulate the pH changes of egg white during the incubation, the course of the conversion into S-ovalbumin was very similar to that either in fertile or unfertile eggs. Therefore, we conclude that thermostabilization of ovalbumin in fertile eggs proceeds by a certain mechanism which depends on the alkalinity of egg white.

Animals↗