Seroprevalence of Toxocara canis infection among children in Swaziland, southern Africa.

As there appeared to be no data available on Toxocara canis infection in the children of Swaziland, a serological survey of T. canis infection was recently conducted among 92 children aged 3-12 years from rural slums in the low- and middle-veld. A child was considered seropositive if, in western blots based on the excretory-secretory antigens of larval T. canis, his or her serum gave a positive result when diluted 1 : 64. Forty-one (44.6%) of the children were found seropositive. There were no statistically significant differences in seroprevalence between the 49 boys and 43 girls investigated (46.9% v. 41.8%) or between the eight subjects aged 12 years and the 47 aged < or = 5 years (62.5% v. 38.3%); the corresponding odds ratios were 0.81 (95% confidence interval=0.36-1.86; P=0.62) and 2.69 (95% confidence interval=0.57-12.62; P=0.20), respectively. The 66 subjects from the middleveld were, however, significantly more likely to be seropositive than the 26 subjects from the lowveld (54.5% v. 19.2%; odds ratio=5.04, with a 95% confidence interval of 1.70-14.98; P<0.01). It seems likely that T. canis infection is common among the children who live in slums in Swaziland, particularly in the country's middleveld, probably as the result of poor hygiene and poor sanitation.

Increased sensitivity to sparsely ionizing radiation due to excessive base excision in clustered DNA damage sites in Escherichia coli.

PURPOSE: In order to clarify the cellular processing and repair mechanisms for radiation-induced clustered DNA damage, we examined the correlation between the levels of DNA glycosylases and the sensitivity to ionizing radiation in Escherichia coli. MATERIALS AND METHODS: The lethal effects of gamma-rays, X-rays, alpha-particles and H2O2 were determined in E. coli with different levels of DNA glycosylases. The formation of double-strand breaks by post-irradiation treatment with DNA glycosylase was assayed with gamma-irradiated plasmid DNA in vitro. RESULTS: An E. coli mutM nth nei triple mutant was less sensitive to the lethal effect of sparsely ionizing radiation (gamma-rays and X-rays) than the wild-type strain. Overproduction of MutM (8-oxoguanine-DNA glycosylase), Nth (endonuclease III) and Nei (endonulease VIII) increased the sensitivity to gamma-rays, whereas it did not affect the sensitivity to alpha-particles. Increased sensitivity to gamma-rays also occurred in E. coli overproducing human 8-oxoguanine-DNA glycosylase (hOgg1). Treatment of gamma-irradiated plasmid DNA with purified MutM converted the covalently closed circular to the linear form of the DNA. On the other hand, overproduction of MutM conferred resistance to H2O2 on the E. coli mutM nth nei mutant. CONCLUSIONS: The levels of DNA glycosylases affect the sensitivity of E. coli to gamma-rays and X-rays. Excessive excision by DNA glycosylases converts nearly opposite base damage in clustered DNA damage to double-strand breaks, which are potentially lethal.

Strong static magnetic field and the induction of mutations through elevated production of reactive oxygen species in Escherichia coli soxR.

PURPOSE: Although strong static magnetic fields (SMF) are supposed to have the potential to affect biological systems, the effects have not been evaluated sufficiently. Experiments should be performed with a powerful SMF-generating apparatus to evaluate the biological effects of SMF. MATERIALS AND METHODS: An Escherichia coli mutation assay was used to assess the mutagenic effects of strong SMF. Various mutant strains of E. coli were exposed to up to 9 Tesla (T) for 24 h and the frequencies of rifampicin-resistant mutations were then determined. The expression of the soxS::lacZ fusion gene was assessed by measurement of beta-galactosidase activity. RESULTS: The results for survival or mutation were obtained with wild-type E. coli strain GC4468 and its derivatives defective in DNA repair enzymes or redox-regulating enzymes were all negative. On the other hand, the mutation frequency was significantly increased by the SMF exposure in soxR and sodAsodB mutants, which are defective in defence mechanisms against oxidative stress. Furthermore, the expression of superoxide-inducible soxS::lacZ fusion gene was stimulated 1.4- and 1.8-fold in E. coli when exposed to 5 and 9 T, respectively. CONCLUSIONS: These results indicate that strong SMF induce mutations through elevated production of intracellular superoxide radicals in E. coli.

Inhibition of beta 2 integrin receptor and Syk kinase signaling in monocytes by the Src family kinase Fgr.

While beta 2 integrin ligand-receptor recognition interactions are well characterized, less is known about how these events trigger signal transduction cascades to regulate the transition from tethering to firm adhesion, spreading, and transendothelial migration. We have identified critical positive and negative regulatory components of this cascade in monocytes. Whereas the Syk tyrosine kinase is essential for beta 2 integrin signaling and cell spreading, the Src family kinase Fgr is a negative regulator of this pathway. Fgr selectively inhibits beta 2 but not beta 1 integrin signaling and Syk kinase function via a direct association between the Fgr SH2 domain and Syk tyrosine Y342. The inhibitory effects of Fgr are independent of its kinase activity, are dose dependent, and can be overcome by chemokines and inflammatory mediators.

Negative regulation of phagocytosis in murine macrophages by the Src kinase family member, Fgr.

Ingestion of opsonized pathogens by professional phagocytes results in the generation and release of microbicidal products that are essential for normal host defense. Because these products can result in significant tissue injury, phagocytosis must be regulated to limit damage to the host while allowing for optimal clearance and destruction of opsonized pathogens. To pursue negative regulation of phagocytosis, we assessed the effect of the Src kinase family member, Fgr, on opsonin-dependent phagocytosis by mouse macrophages. We chose Fgr because it is present in high concentrations in circulating phagocytes but is not essential for Fcgamma receptor-mediated ingestion by mouse macrophages. Although expression of Fgr both in a macrophage cell line and in primary macrophages significantly attenuates ingestion mediated by Fcgamma receptors and CR3, it does not affect macropinocytosis or receptor-mediated endocytosis. This selective effect of Fgr is independent of its tyrosine kinase function. After Fcgamma receptor cross-linking, Fgr becomes associated with the immunoreceptor tyrosine-based inhibition motif (ITIM)-containing receptor, SIRPalpha (a member of the signal-regulatory protein family, also known as Src homology 2 domain-containing protein tyrosine phosphatase [SHP] substrate 1 [SHPS-1], brain immunoglobulin-like molecule with tyrosine-based activation motifs [BIT], and P84) and potentiates the association of the phosphatase SHP-1 with SIRPalpha. This association is responsible, at least in part, for decreasing positive signaling essential for optimal phagocytosis. These data demonstrate an important negative regulatory role for this Src kinase family member and suggest that this homeostatic function must be overcome for optimal uptake and clearance of opsonized pathogens.

Isolation, characterization, and chromosomal mapping of a novel cDNA clone encoding human selenium binding protein.

We have isolated the full-length human 56 kDa selenium binding protein (hSP56) cDNA clone, which is the human homolog of mouse 56 kDa selenium binding protein. The cDNA is 1,668 bp long and has an open reading frame encoding 472 amino acids. The calculated molecular weight is 52.25 kDa and the estimated isoelectric point is 6.13. Using Northern blot hybridization, we found that this 56 kDa selenium binding protein is expressed in mouse heart with an intermediate level between those found in liver/lung/kidney and intestine. We have also successfully expressed hSP56 in Escherichia coli using the expression vector-pAED4. The hSP56 gene is located at human chromosome 1q21-22).

Protein pI alteration related to strain variation of infectious bronchitis virus, an avian Coronavirus.

Viral proteins of two strains of infectious bronchitis virus (IBV), which have different tissue trophism and serology, were separated on the basis of their isoelectric points (pI). The viruses have four structural proteins; the protein of greatest serological importance is found at the peplomer tip. The viral structural proteins separated by isoelectric focusing were identified by comparison to SDS-PAGE separations. Three protein bands were identical in pI and one protein band showed a difference in pI between strains. When the renatured viral proteins were Western blotted and reacted with strain-specific antiserum, antigen-antibody complexing was seen only at points corresponding to the strain-specific variant bands. For IBV strain Mass-41, antigen-antibody complexing occurred at a pI of 6.8, and, for IBV strain Ark-99, at 7.2. No cross reaction of antisera was observed for either strain. Since tissue affinities are a function of the viral peplomer-mediated attachment of virus to cells and are often directly related to pathogenicity, it appears that altered pathogenicity of strains of IBV may be detected by alteration of pI of the proteins. Classification by pI of proteins of at least the smaller viruses allows untypeable, highly pathogenic or persistent strains of these viruses to be characterized on the basis of variant proteins.

Detection of antibody to avian viruses in human populations.

The ability of three avian viruses to elicit antibody response in humans was surveyed for the purpose of identifying zoonotic diseases. Antibody levels in people associated with poultry were compared to those in people having limited poultry association. Antibody levels to three avian viruses: infectious bursal disease virus, a birnavirus; Newcastle disease virus, a paramyxovirus; and avian infectious bronchitis virus, a coronavirus were determined by enzyme-linked immunosorbent assays (ELISA). Differences between the two study groups were evident: people having a known association with poultry showed significantly higher levels of antibodies to Newcastle disease and avian infectious bronchitis virus. Antibodies detected may be due to virus exposure rather than zoonoses.

Physiological effects of adrenocorticotropic hormone and hydrocortisone in laying hens.

To study the hormonal effects on hematologic parameters as indicators of chronic stress, exogenous adrenocorticotropin (ACTH) at 6.3 or 20.0 IU/kg/day and hydrocortisone at .25 or 2.5 mg/kg/day were administered parenterally to laying hens. Both ACTH treatments induced significant (P less than .05) heterophilia, monocytosis, eosinophilia, and basophilia. Significantly elevated leucocyte counts and lymphopenia (P less than .05) were observed with the high dosage of ACTH. Both hydrocortisone-treated groups developed an absolute lymphopenia and heterophilia (P less than .05). The low dosage of hydrocortisone induced a significant (P less than .05) monocytosis; the high dosage caused significant (P less than .05) decreases in the total eosinophil and basophil counts as well as an increase in the ratio of heterophils to lymphocytes. The hemopoietic parameters, especially heterophil counts, were sensitive indicators of a hormonal stress response induced by the administration of ACTH and hydrocortisone.

Detection of latent avian adenovirus-associated virus proteins in chicken cells.

Chicken kidney cells, derived from the eggs of white leghorn chickens that had serological evidence of prior exposure to both adenovirus and the adeno-associated virus (AAV), produced AAV antigenic proteins upon challenge with purified adenovirus. Antigen was detected by indirect immunofluorescence using monoclonal antibody to AAV. The number of fluorescent cells were few and did not increase during the course of adenovirus infection. Similar results were obtained using cells prepared from specific-pathogen-free chicks with no previous exposure to adenovirus or AAV. It is postulated that the avian AAV can exist as a latent infection in the germ line of chickens.

Use of egg yolk to determine antibody levels in chickens inoculated with a hemagglutinating duck adenovirus (adenovirus 127-like).

Chickens were experimentally infected with a duck adenovirus that has been shown to be serologically indistinguishable from Adenovirus 127. Sera and eggs were collected at intervals after exposure for antibody determination by the hemagglutination-inhibition (HI) test, the enzyme-linked immunosorbent assay (ELISA), and the immunodiffusion (ID) test. Egg yolks were processed for use in the serological tests by (a) dilution in phosphate-buffered saline (PBS), (b) extraction of the water-soluble fraction with chloroform, or (c) freezing and thawing PBS-diluted yolks and testing the supernatant fluid. HI antibody titers from serum and extracted yolk were similar except during the initial 2 weeks, when yolk antibody levels were low or absent. Chloroform-extracted yolks were suitable material for the HI, ELISA, and ID tests. Heat inactivation of the chloroform-extracted yolk had no effect on titers.

Morphogenesis of canary poxvirus and its entrance into inclusion bodies.

A virus isolated from a natural outbreak of canarypox was replicated on the chorioallantoic membranes of chicken embryos, and its ultrastructure and development were observed. Electron microscopy of thin sections of pocks produced on the chorioallantoic membranes revealed a variety of developmental forms which appear similar to those demonstrated in studies of vaccinia, ie, viroplasm or viral factories; immature, undifferentiated virions partially enclosed by membranes; completely enclosed nondifferentiated spherical or oval virions; immature virions with discrete nucleoids; and the more compact brick-shaped mature virions. Two types of A-type inclusions were noted: those with virions around the periphery, and those filled with virus particles. The appearance of mature viruses within the inclusion bodies and different stages of viruses outside the inclusion indicate that in a course of development, maturing poxvirus may enter the inclusion bodies as they acquire surface tubules on their envelopes. Mature virions also were seen budding out of the cell membrane, apparently enveloped in a portion of the membrane. Studies showing the entrance of poxvirus into inclusion bodies have not been reported. In this report, electron micrographs are shown of viruses entering inclusion bodies.

Use of egg yolk in serological tests (ELISA and HI) to detect antibody to Newcastle disease, infectious bronchitis, and Mycoplasma gallisepticum.

Serum and yolks from commercial flocks and from hens exposed to Newcastle disease virus (NDV), infectious bronchitis virus (IBV), and Mycoplasma gallisepticum (MG) were tested for immunoglobulin G antibody by the enzyme-linked immunosorbent assay (ELISA) and the hemagglutination-inhibition (HI) test. Yolks prepared by chloroform extraction and low-speed centrifugation performed well in the serological tests used and were a suitable alternative to serum for antibody determination by the ELISA for NDV, IBV, and MG and by HI test for NDV.

Selection of bacteria with favorable transport properties through porous rock for the application of microbial-enhanced oil recovery.

This paper presents a bench-scale study on the transport in highly permeable porous rock of three bacterial species-Bacillus subtilis, Pseudomonas putida, and Clostridium acetobutylicum-potentially applicable in microbial-enhanced oil recovery processes. The transport of cells during the injection of bacterial suspension and nutrient medium was simulated by a deep bed filtration model. Deep bed filtration coefficients and the maximum capacity of cells in porous rock were measured. Low to intermediate ( approximately 10/ml) injection concentrations of cellular suspensions are recommended because plugging of inlet surface is less likely to occur. In addition to their resistance to adverse environments, spores of clostridia are strongly recommended for use in microbial-enhanced oil recovery processes since they are easiest among the species tested to push through porous rock. After injection, further transport of bacteria during incubation can occur by growth and mobility through the stagnant nutrient medium which fills the porous rock. We have developed an apparatus to study the migration of bacteria through a Berea sandstone core containing nutrient medium.

Is avian adeno-associated virus an endogenous virus of chicken cells?

The adeno-associated viruses (AAV) are defective parvoviruses which produce infective progeny only in cells co-infected with a 'helper' adenovirus (Ad). Both human and simian AAV have been recovered from human and simian primary cell cultures following their inoculation with 'AAV-free' Ad. Whereas some studies have suggested that AAV exists in a latent state in these cells, others have indicated that the AAV genome is capable of establishing and maintaining a latent state in defined laboratory conditions which mimic the situation proposed for the 'latent' AAV recovered from human and simian tissues. Here, avian adeno-associated virus (AAAV) was consistently recovered from limiting dilutions of purified and unpurified avian Ad stocks propagated in embryonating chicken eggs derived from two independently raised flocks of White Leghorn (WL) chickens but not when these Ad stocks were propagated in duck cells. These observations suggest that AAAV is a latent endogenous virus of at least some flocks of WL chickens.

The effect of incubation temperature on the propagation of duck adenovirus (virus 127-like) in duck and chicken cells.

Duck adenovirus (Cornell strain) was propagated in duck and chicken embryo cells at 37.5 C and at 40 C. In duck cells, virus levels, as indicated by HA titers, peaked earlier at 40 C than at 37.5 C. High titers were eventually observed in duck cells at both temperatures. In chicken embryo fibroblasts, no titers were observed at 37.5 C, whereas low titers were observed at 40 C. Evidence of virus propagation was not detected in chicken embryo liver and kidney cells.

Neutralizing antibodies to CELO and avian adenovirus-associated viruses in the albumen of chicken eggs.

Neutralizing antibodies to CELO virus and to avian adenovirus-associated virus (A-AV) were detected in the albumen of eggs from four hens inoculated with these viruses. The antibody concentrations of serum, yolk, and albumen were determined before inoculation and at various times postinoculation (PI) by enzyme-linked immunosorbent assay (ELISA) and virus-neutralization (VN) tests. The antibody concentration in albumen was 0.3% to 1.0% of that detected in serum and yolk. Uninoculated hens showed no detectable antibody in serum, yolk, or albumen. It is suggested that the presence of antibody in the egg albumen may play a role in egg-transmission of viruses.

Enhancement and inhibition of CELO virus pathogenicity in quail by avian adenovirus-associated virus.

Dual infection of 12 day-old quail (Colinus virginianus) with 10(6) plaque forming units of CELO virus and low doses of avian adeno-associated virus (A-AV), resulted in significant enhancement of CELO virus-induced mortality, whereas dual infections with high doses of A-AV resulted in a delay in mortality. A-AV induced enhancement and inhibition of CELO virus pathogenicity could be blocked by the addition of A-AV antiserum prior to infection.