[Cytochemical research on the reproduction of tobacco mosaic virus. I. UV spectrophotometric measurement of an isolated viral substance].
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Research↗
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[Staphylococcal syndrome of burned skin: the isolation of II-phage group Staphylococcus aureus and the experimental reproduction of the syndrome].
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Animals↗
Sperm preparation for ART.
The onset of clinical assisted reproduction, a quarter of a century ago, required the isolation of motile spermatozoa. As the indication of assisted reproduction shifted from mere gynaecological indications to andrological indications during the years, this urged andrological research to understand the physiology of male germ cell better and develop more sophisticated techniques to separate functional spermatozoa from those that are immotile, have poor morphology or are not capable to fertilize oocytes. Initially, starting from simple washing of spermatozoa, separation techniques, based on different principles like migration, filtration or density gradient centrifugation evolved. The most simple and cheapest is the conventional swim-up procedure. A more sophisticated and most gentle migration method is migration-sedimentation. However, its yield is relatively small and the technique is therefore normally only limited to ejaculates with a high number of motile spermatozoa. Recently, however, the method was also successfully used to isolate spermatozoa for intracytoplasmic sperm injection (ICSI). Sperm separation methods that yield a higher number of motile spermatozoa are glass wool filtration or density gradient centrifugation with different media. Since Percoll as a density medium was removed from the market in 1996 for clinical use in the human because of its risk of contamination with endotoxins, other media like IxaPrep, Nycodenz, SilSelect, PureSperm or Isolate were developed in order to replace Percoll. Today, an array of different methods is available and the selection depends on the quality of the ejaculates, which also includes production of reactive oxygen species (ROS) by spermatozoa and leukocytes. Ejaculates with ROS production should not be separated by means of conventional swim-up, as this can severely damage the spermatozoa. In order to protect the male germ cells from the influence of ROS and to stimulate their motility to increase the yield, a number of substances can be added to the ejaculate or the separation medium. Caffeine, pentoxifylline and 2-deoxyadenosine are substances that were used to stimulate motility. Recent approaches to stimulate spermatozoa include bicarbonate, metal chelators or platelet-activating factor (PAF). While the use of PAF already resulted in pregnancies in intrauterine insemination, the suitability of the other substances for the clinical use still needs to be tested. Finally, the isolation of functional spermatozoa from highly viscous ejaculates is a special challenge and can be performed enzymatically to liquefy the ejaculate. The older method, by which the ejaculate is forcefully aspirated through a narrow-gauge needle, should be abandoned as it can severely damage spermatozoa, thus resulting in immotile sperm.
Cell Separation↗
Molecular characterization of the GnRH system in zebrafish (Danio rerio): cloning of chicken GnRH-II, adult brain expression patterns and pituitary content of salmon GnRH and chicken GnRH-II.
The zebrafish has proven to be a model system with unparalleled utility in vertebrate genetic and developmental studies. Substantially less attention has been paid to the potential role that zebrafish can play in answering important questions of vertebrate reproductive endocrinology. As an initial step towards exploiting the advantages that the zebrafish model offers, we have characterized their gonadotropin-releasing hormone (GnRH) system at the molecular level. GnRHs comprise a family of highly conserved decapeptide neurohormones widely recognized to orchestrate the hormonal control of reproduction in all vertebrates. We have isolated the gene and cDNA encoding chicken GnRH-II (cGnRH-II) from zebrafish, as well as several kilobases of upstream promoter sequence for this gene. As the gene encoding salmon GnRH (sGnRH) has been previously isolated (Torgersen et al, 2002), this is the second GnRH gene isolated from zebrafish to date. We have localized expression of these two genes in the brains of reproductively mature zebrafish using in situ hybridization. sGnRH is localized to the olfactory bulb-terminal nerve region (OB-TN), the ventral telencephalon-preoptic area (VT-POA) and, as we report here for the first time in any teleost species, the hindbrain. cGnRH-II is expressed exclusively in the midbrain, as has been found in all other jawed vertebrate species examined. Finally, the levels of both GnRH peptides in pituitaries of reproductively mature zebrafish were quantified using specific ELISAs. sGnRH pituitary peptide levels were shown to be 3- to 4-fold higher than cGnRH-II pituitary peptide. The cumulative results of these experiments allow us to conclude that zebrafish express just two forms of GnRH in a site-specific manner within the brain, and that sGnRH is the hypophysiotropic GnRH form. This work lays the foundation for further research into the control of reproduction in zebrafish, such as the functional significance of multiple GnRHs in vertebrates, and the molecular mechanisms controlling tissue-specific GnRH expression.
Animals↗
Isolation and partial characterization of a fatty-acid-binding protein from Ascaris suum reproductive tissue.
A 12-kDa fatty-acid-binding protein was purified to homogeneity from Ascaris suum reproductive tissue as confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. N-terminal amino-acid-sequence analysis of the protein revealed its identity with the ABA-1 allergen protein isolated from A. suum pseudocoelomic fluid. Fatty-acid binding by the protein from A. suum reproductive tissue was investigated using the Lipidex 1000 assay, which revealed the presence of a single class of fatty-acid-binding sites with an apparent dissociation constant for palmitate of about 0.8 microM.
Allergens↗
Identification of two isolates of Trichinella recovered from humans in France.
Two Trichinella isolates from humans in France were characterized using reproductive capacity indices and a combination of molecular methods. The isolate TRLL hybridized with the pig type-specific probe pPra and had pig type restriction profiles and rDNA patterns. It was therefore identified as a domestic or pig type isolate. The isolate CTRD-85 had similarities and differences in restriction profiles and rDNA patterns with both AF1 and Trichinella nelsoni and was identified as a sylvatic type. Pattern comparisons also show that T. nelsoni is similar to variants of the North American sylvatic type.
Animals↗
A comparison of three geographical isolates of Trichinella spiralis from the mid-Atlantic United States.
Three isolates of Trichinella spiralis (black bear: 41o50'N, 79o00'W, 1982; grey fox: 41o00'N, 76o00'W, 1982; domestic pig: 39o80'N, 75o30'W, 1983) from the mid-Atlantic United States were characterized in Crl: COBS CFW (SW) mice during 10 successive 40-day generations. Over 10 generations, the isolates differed as follows: worm position of the ursine isolate was significantly more posteriad compared to the porcine isolate; sex ratio of the vulpine isolate was significantly lower compared to the ursine and porcine isolates; females of the porcine isolate produced significantly more newborn larvae in vitro than the sylvatic isolates; both the larvae per gram (LPG) and reproductive capacity index (RCI) were significantly higher for the porcine isolate; and male worms of the vulpine isolate were significantly smaller than those of the porcine or ursine isolates. No differences were observed among isolates for percentage of inoculum recovered, length of female worms, or uterine length. The fecundity of the isolates in hamsters, gerbils, multimammate rats, Sprague-Dawley rats, and Peromyscus sp. was examined also. Based upon RCI and LPG, the porcine isolate was the most fecund in all hosts except gerbils. This higher fecundity was often manifest as increased morbidity and mortality in hosts infected with this isolate. The relationship between the virulent nature of the porcine isolate and the enhanced transmission of T. spiralis are discussed.
Animals↗
Temporal characterization of transplacental infection of porcine fetuses with porcine reproductive and respiratory syndrome virus.
Pregnant gilts were exposed to porcine reproductive and respiratory syndrome virus (PRRSV) by IV inoculation at or about gestation day 30 (3 gilts), 50 (3 gilts), 70 (3 gilts), or 90 (5 gilts) to investigate the likelihood of transplacental infection with PRRSV at various stages of gestation. At or about 3, 6, and 9 weeks after exposure, gilts were either euthanatized while still pregnant or allowed to farrow. Gilts and pigs were observed for clinical signs of infection, and gilts, pigs, and fetuses were tested for PRRSV and homologous antibody. All gilts were healthy throughout the study, except that farrowing was sometimes difficult and prolonged, and 2 gilts failed to farrow the entire litter. One gilt farrowed on day 111 of gestation; all others farrowed on day 114 or later. Porcine reproductive and respiratory virus was isolated from significantly (chi 2 test, P < 0.01) more fetuses and live and stillborn pigs of the 5 gilts that were infected at 90 or 92 days of gestation than from the fetuses and live and stillborn pigs of the 9 gilts that were infected at 72 or fewer days of gestation (ie, 33 of 44, 75% vs 3 of 78, 4%). After initial infection, PRRSV was isolated from gilts and their pigs for a maximum of 3 weeks and 8 to 11 weeks, respectively. Findings of this study, with regard to the temporal aspects of transplacental infection, may help explain why natural epizootics of PRRSV-induced maternal reproductive failure are often recognized principally as problems of late-term gestation and neonatal survival.
Animals↗
Temporal localization of porcine reproductive and respiratory syndrome virus in reproductive tissues of experimentally infected boars.
Porcine reproductive and respiratory syndrome virus (PRRSV) has been reported to be shed in the semen of infected boars. To determine whether the reproductive tissues could be a persistent source of virus and the possible origin of PRRSV found in semen of infected boars, 20 PRRSV-seronegative boars were intranasally inoculated with 5 x 10(6) median tissue culture infective doses (TCID50) of PRRSV and necropsied at different times post-inoculation (p.i.) from Day 2 to Day 37 p.i. Blood samples were collected before experimental inoculation, at necropsy and at different times p.i. At necropsy, epididymal semen and reproductive tissues were collected and the presence of the virus determined by virus isolation. The infection of the boars was demonstrated by the isolation of the virus from the sera of all inoculated boars and by seroconversion. PRRSV was detected in serum samples from Day 2 to Day 23 p.i., although the viremic period was largely dependent on the individual response to infection. Viral replication was proven within different reproductive tissues from Day 2 to Day 23 p.i., being most consistently found in the epididymus. In addition, PRRSV was isolated in semen from Day 4 to Day 10 p.i. The correlation of a diminished viremia and the inability to isolate PRRSV from semen or reproductive tissues may be due to one of two possibilities. First, viremia is responsible for most of the virus isolated from reproductive tissues due to the movement of PRRSV-infected cells out of the blood and into the tissues. Second, viremia may initially seed the reproductive tissues with PRRSV, and then the virus is produced into the reproductive tract and shed into semen at low levels.
Animals↗
Model parameters and outbreak control for SARS.
Control of the 2002-2003 severe acute respiratory syndrome (SARS) outbreak was based on rapid diagnosis coupled with effective patient isolation. We used uncertainty and sensitivity analysis of the basic reproductive number R0 to assess the role that model parameters play in outbreak control. The transmission rate and isolation effectiveness have the largest fractional effect on R0. We estimated the distribution of the reproductive number R0 under perfect isolation conditions. The distribution lies in the interquartile range 0.19-1.08, with a median of 0.49. Even though the median of R0 is <1, we found that 25% of our R0 distribution lies at R0 > 1, even with perfect isolation. This implies the need to simultaneously apply more than one method of control.
Communicable Disease Control↗
Current trends in reproductive immunology: an overview.
Over the past decade has come general recognition that, directly or indirectly, immunology intrudes into nearly every aspect of mammalian reproduction. Charles Darwin's notion that the profligacy of women led to reduced fertility gained plausibility with the subsequent discovery of the auto- and alloantigenicity of spermatozoa and of testicular material. From this observation arose the reasonable expectation that immunological control of fertility may be feasible. Discovery of the importance of natural transfer of immunity from mother to offspring, the ontogeny of the immune response, and recognition that pregnancy is an almost consistently successful violation of the "laws of transplantation' are only a few highlights or components of the burgeoning, multifaceted field we have come to recognize as the immunology of reproduction. An overview of this subject is here presented with regard to its evolutionary origins, its accomplishments, its current trends, and some of its potentialities. The immunology of reproduction has not developed in isolation; in recent years it has benefited enormously from developments in other fields, and in its turn it has exerted its own impact on other disciplines, especially on transplantation. The present preoccupation of many immunologists with immunoregulation stems largely from independent discoveries in the realm of reproductive immunobiology - the etiology of Rhesus disease and its prophylaxis, and the principle of immunological tolerance, from investigations on the peculiarities of dizygotic twins in cattle.
Antibodies↗
Immunology of reproduction: present trends.
Over the past decade has come general recognition that, directly or indirectly, immunology intrudes into nearly every aspect of mammalian reproduction. Charles Darwin's notion that the profligacy of women led to reduced fertility gained plausibility with the subsequent discovery of the auto- and alloantigenicity of spermatozoa and of testicular material. From this observation arose the reasonable expectation that immunological control of fertility may be feasible. Discovery of the importance of natural transfer of immunity from mother to offspring, the ontogeny of the immune response and recognition that pregnancy is an almost consistently successful violation of the "laws of transplantation" are only a few highlights or components of the burgeoning, multifaceted field we have come to recognize as the immunology of reproduction. An overview of this subject will be presented with regard to its evolutionary origins, its accomplishments, its current trends and some of its potentialities. The immunology of reproduction has not developed in isolation; in recent years it has benefited enormously from developments in other fields, and in its turn it has exerted its own impact on other disciplines, especially on transplantation. The present preoccupation of many immunologists with immunoregulation stems largely from independent discoveries in the realm of reproductive immunobiology: the etiology of Rh disease and its prophylaxis, and the principle of immunological tolerance, from investigations on the peculiarities of dizygotic twins in cattle.
Allergy and Immunology↗
Plant polyphenolic complex inhibits the reproduction of influenza and herpes simplex viruses.
A polyphenolic complex (PC), isolated from the Bulgarian medicinal plant Germanium sanguineum L., inhibited the reproduction of influenza virus types A and B in vitro and in ovo. It protected white mice in an experimental lethal influenza infection. When influenza viruses were treated with a high dose (1 mg/mL), their hemagglutination, neuraminidase and infections activities were reduced completely. It was assumed that the PC-specific inhibitory effect on influenza virus reproduction in tissue cultures (50 micrograms/mL) was mediated by partial inactivation of viral glycoproteins which prevented the initial stages of reproduction. The PC also affected the intracellular stages of viral replication. The effect was dependent on the mode of application, the viral strain, and the biological test system. The PC also inhibited the reproduction of herpes simplex virus in vitro. Infectious titers were reduced and newly synthesized virion with damaged protein envelopes were observed by electron microscopy.
Animals↗
Metabolic and hormonal control of the desire for food and sex: implications for obesity and eating disorders.
During evolution, the ability to overeat and store the extra energy as glycogen and lipids in specialized tissues must have conferred a reproductive advantage by releasing animals from the need to eat constantly, enabling them to engage in behaviors that improved reproductive success. Mechanisms that inhibited ingestive behavior might have been most adaptive when they caused individuals to stop foraging, hoarding and eating in order to find and court potential mates. Conversely, the ability to abstain from reproductive activities to engage in foraging and eating was probably critical for individual survival during severe energetic challenges because reproductive processes are energetically costly and can be delayed until the energetic conditions improve. The mechanisms that control ingestive behavior most likely evolved under conditions in which both food and mates were available, and thus, our understanding might be limited by our narrow focus on food intake in animals isolated from potential mates, and reproductive behaviors in the absence of food. Our understanding of obesity and eating disorders will be enriched by the study of the choice between ingestive and reproductive behaviors and by a renewed attention to "reproductive" hormones such as gonadal steroids and hypothalamic releasing hormones. Furthermore, leptin and reproductive hormones have both organizational and activational effects on the energy balancing system including those mechanisms that control appetite, body fat content and body fat distribution. Understanding these organizational and activational effects on body fat distribution might lead to a better understanding of sex differences in the propensity to develop obesity, type II diabetes and eating disorders.
Animals↗
Expression and topographical localization of cell surface fucosyltransferase activity during epididymal sperm maturation in the mouse.
We have demonstrated previously that spermatogenic cells in the mouse testis have high levels of fucosyltransferase activity. Furthermore, a significant portion of this activity has been localized to the cell surface (Millette et al.: Cell Biology of the Testis and Epididymis, 1987). Differential expression of fucosyltransferases and their function as ecto-enzymes may be important in the processes of sperm maturation and fertilization in mammals. Accordingly, here we report the activity levels of fucosyltransferase (FT) in spermatozoa isolated from the mouse caput and cauda epididymides. Calculated on a per cell basis, spermatozoa from the caput epididymis have significantly more FT activity than do spermatozoa from the cauda epididymis (18.07 +/- 2.2 pmol/million cells compared with 2.8 +/- 0.09 pmol/million cells). Furthermore, caput sperm exhibit a more significant increase in FT activity when assayed in the presence of Nonidet P-40. Calculated on the basis of cell surface area, however, FT activity remains constant on the head portion of spermatozoa isolated from all portions of the male reproductive tract and from capacitated spermatozoa. Measurements of FT activity in extracts of isolated sperm tails from cells at different stages of maturation indicate a greatly diminished activity in tails from sperm in the cauda epididymis. The total sperm surface area is composed predominantly of the plasma membrane surrounding the flagellar apparatus. Therefore, our data demonstrate that FT activity is retained selectively on the different topological regions of sperm, with losses during sperm maturation in the epididymis being restricted to the tail segment. Maintenance of high levels of FT activity of the plasma membranes of the mouse sperm head raise the possibility that FT is indeed involved in some aspects of sperm-egg recognition.
Animals↗
[The role of adrenergic receptors in the regulation of blood flow in the reproductive organs of swine during the estrous cycle].
The dimension of flow in uterine artery (ua) and ovarian artery (oa) was measured. It was conducted in the isolated porcine (100-130 kg/per head) reproductive organs perfused with their own blood or Krebs-Henseleit fluid when the pressure was kept on the constant level 100 m Hg. Phenoxybenzamine (Ph), phentolamine (R), propranolol (P), pronetalol (Pr) were used and also after them adrenaline (A), noradrenaline (NA) or isoprenaline (I) were administrated. Changes in blood flow (bf) through the organs connected with inhibition or stimulation of adrenergic receptors during estrous cycle were analysed. It was stated that R increased bf, but not significantly. Ph increased significantly bf in the reproductive organs and blocked decreasing bf influence of A and NA in ua and oa areas. Both A and I administrated after Ph and R caused the increase in bf in the both investigated vessel areas. Beta-adrenolytic agents P and Pr decreased bf and also blocked increasing bf action of I. When we used A or NA after P or Pr administration, there was observed significant limitation of bf in the reproductive organs. Reactions evoked by alpha- and beta-adrenergic stimulation were different during estrous cycle. The highest activity of alpha receptor--which dominates the vessels of porcine reproductive organs--was found in the luteal phase of cycle where the activity of beta receptor was the lowest. The activity of alpha receptor decreased and increased activity of beta receptor in the pre and postovulatory phase of cycle compared to the state observed in the luteal phase. Beta receptor did not play any significant role in the regulation of bf in the porcine reproductive organs. Data from references discussed in this work and our results suggest that regulation of sensitivity of vessels in the porcine reproductive organs was not connected with quantitative representation of adrenergic receptors. Changes of vessel sensitivity were connected with changes of alpha-adrenergic receptor activity.
Animals↗
Comparison of molecular and biological characteristics of a modified live porcine reproductive and respiratory syndrome virus (PRRSV) vaccine (ingelvac PRRS MLV), the parent strain of the vaccine (ATCC VR2332), ATCC VR2385, and two recent field isolates of PRRSV.
The objectives of this study were to compare the molecular and biological characteristics of recent porcine reproductive and respiratory syndrome virus (PRRSV) field isolates to those of a modified live virus (MLV) PRRS vaccine and its parent strain. One hundred seventeen, 4-week-old pigs were randomly assigned to six groups. Group 1 (n = 20) served as sham-inoculated negative controls, group 2 (n = 19) was inoculated with Ingelvac PRRS MLV vaccine, group 3 (n = 20) was inoculated with the parent strain of the vaccine (ATCC VR2332), group 4 (n = 19) was inoculated with vaccine-like PRRSV field isolate 98-38803, group 5 (n = 19) was inoculated with PRRSV field isolate 98-37120, and group 6 (n = 20) was inoculated with known high-virulence PRRSV isolate ATCC VR2385. The levels of severity of gross lung lesions (0 to 100%) among the groups were significantly different at both 10 (P < 0.0001) and 28 days postinoculation (p.i.) (P = 0.002). At 10 days p.i., VR2332 (26.5% +/- 4.64%) and VR2385 (36.4% +/- 6.51%) induced gross lesions of significantly greater severity than 98-38803 (0.0% +/- 0.0%), 98-37120 (0.8% +/- 0.42%), Ingelvac PRRS MLV (0.9% +/- 0.46%), and negative controls (2.3% +/- 1.26%). At 28 days p.i., 98-37120 (17.2% +/- 6.51%) induced gross lesions of significantly greater severity than any of the other viruses. Analyses of the microscopic-interstitial-pneumonia-lesion scores (0 to 6) revealed that VR2332 (2.9 +/- 0.23) and VR2385 (3.1 +/- 0.35) induced significantly more severe lesions at 10 days p.i. At 28 days p.i., VR2385 (2.5 +/- 0.27), VR2332 (2.3 +/- 0.21), 98-38803 (2.6 +/- 0.29), and 98-37120 (3.0 +/- 0.41) induced significantly more severe lesions than Ingelvac PRRS MLV (0.7 +/- 0.17) and controls (0.7 +/- 0.15). The molecular analyses and biological characterizations suggest that the vaccine-like isolate 98-38803 (99.5% amino acid homology based on the ORF5 gene) induces microscopic pneumonia lesions similar in type to, but different in severity and time of onset from, those observed with virulent strains VR2385 and the parent strain of the vaccine. Our data strongly suggest that isolate 98-38803 is a derivative of Ingelvac PRRS MLV and that the isolate is pneumovirulent.
Animals↗
[Antiviral factor isolated from infected cell cultures].
An antiviral factor of protein nature inhibiting reproduction of VEE, herpes, vaccinia VSV, fowl plague viruses was isolated from infected cell cultures. The factor has no virucidal or prophylactic effect, stable to low pH values and heating at 100 degrees C for 30 min.
Animals↗