The effect of parenteral alimentation fluid,undiluted or diluted with saline or frseh sera,on the growth of Candida albicans in vitro at 37 °C

Parenteral alimentation is often complicated by Candida albicans infection which may be fatal. This study investigated the effect of alimentation fluid (Aminosol) on C. albicans' growth in vitro. It was found that concentrated Aminosol (1400 milliosmoles) maintained C. albicans in a viable state but inhibited replication. Dilution of alimentation fluid to physiological concentrations (300 milliosmoles) with either saline or aged pooled normal sera promoted in vitro growth of C. albicans which was equivalent to that obtained in BHI broth and was slightly less than that obtained in Sabouraud's broth. The effects of fresh sera with full complement activity were also investigated. In fresh sera appropriately diluted with physiological saline, some clumping of the yeasts was observed and all formed germ tubes. Growth as defined by budding or the formation of hyphae was inhibited. When Aminosol was diluted to 300 milliosmoles with fresh sera, all yeasts were noted to be in clumps with germ tubes as well as continually growing hyphae. Growth was approximately equal to that seen in Aminosol similarly diluted with saline.     

The role of non-specific serum opsonins in thein vitro interaction (adherence) between peritoneal macrophages of newborn precolostral piglets and rough strain ofEscherichia coli

The role of non-antibody, natural opsonins in sera of newborn, precolostral piglets for the early phase of phagocytosis (adhesion) of roughEscherichia coli to peritoneal macrophages of these animals, was studied. Suspensions of macrophages, incubated together with bacteria in the presence or absence of piglet serum opsonins, were submitted to differential centrifugation to enable the separation of macrophage-associated bacteria (i.e. opsonized) from unopsonized, free bacteria in the supernatant. It was found that native piglet sera having no detectable antibody activity toEscherichia coli, do possess significant opsonic activity,i.e. they enhanced thein vitro adherence of roughEscherichia coli to macrophages. Furthermore, this activity could be removed by procedures or substances known to inactivate the complement by different mechanisms: heating for 30 min at 56°C, addition of EDTA, absorption of sera by zymosan, addition of complement inhibitors phosphomannan and carrageenin. These results are interpreted as further evidence for the presence of complement-dependent serum opsonins to roughEscherichia coli in sera of newborn precolostral piglets.     

Plasma-Derived Human C1-Esterase Inhibitor Does Not Prevent Mechanical Ventilation-Induced Pulmonary Complement Activation in a Rat Model of Streptococcus pneumoniae Pneumonia

Mechanical ventilation has the potential to cause lung injury, and the role of complement activation herein is uncertain. We hypothesized that inhibition of the complement cascade by administration of plasma-derived human C1-esterase inhibitor (C1-INH) prevents ventilation-induced pulmonary complement activation, and as such attenuates lung inflammation and lung injury in a rat model of Streptococcus pneumoniae pneumonia. Forty hours after intratracheal challenge with S. pneumoniae causing pneumonia rats were subjected to ventilation with lower tidal volumes and positive end-expiratory pressure (PEEP) or high tidal volumes without PEEP, after an intravenous bolus of C1-INH (200 U/kg) or placebo (saline). After 4 h of ventilation blood, broncho-alveolar lavage fluid and lung tissue were collected. Non-ventilated rats with S. pneumoniae pneumonia served as controls. While ventilation with lower tidal volumes and PEEP slightly amplified pneumonia-induced complement activation in the lungs, ventilation with higher tidal volumes without PEEP augmented local complement activation more strongly. Systemic pre-treatment with C1-INH, however, failed to alter ventilation-induced complement activation with both ventilation strategies. In accordance, lung inflammation and lung injury were not affected by pre-treatment with C1-INH, neither in rats ventilated with lower tidal volumes and PEEP, nor rats ventilated with high tidal volumes without PEEP. Ventilation augments pulmonary complement activation in a rat model of S. pneumoniae pneumonia. Systemic administration of C1-INH, however, does not attenuate ventilation-induced complement activation, lung inflammation, and lung injury.     

Examination of the bactericidal and opsonic activity of normal human serum for a mucoid and nonmucoid strain ofPseudomonas aeruginosa

The bactericidal and opsonic activity of fresh human serum (FHS) for a mucoid strain ofPseudomonas aeruginosa, 144M, and its spontaneous nonmucoid revertant, 144NM, was examined. Strain 144M was sensitive to the bactericidal activity of FHS, but strain 144NM was not. This bactericidal activity was due to the combined interaction of IgG and IgM with complement, activated through both pathways. Neither 144M nor 144NM was ingested by human polymorphonuclear leukocytes (PMNL) without FHS. Whereas maximal phagocytosis of 144M required only 5% FHS, comparable ingestion of 144NM required 25% FHS. Maximal phagocytosis of either 144M or 144NM required IgG, IgM, and complement. However, 144M required a heat-sensitive opsonic IgG, whereas 144NM required a heat-resistant IgG. Using selective absorption techniques, the targets for bactericidal and opsonic immunoglobulins on 144M and 144NM appeared to be different, suggesting that the variant 144NM had one or more altered, absent, or inaccessible cell surface components that account for differences in response to FHS and PMNL.     

The generation of chemotactic activity in sera of germ-free newborn piglets by interaction with rough strain ofEscherichia coli

The interaction of sera of newborn precolostral piglets with rough strain ofEscherichia coli or its endotoxin leads to formation of a factor which is chemotactic for rabbit polymorphonuclear neutrophil leucocytesin vitro (as tested using the Boyden's diffusion two-compartment chamber). Smooth strain ofEscherichia coli does not induce chemotaxin formation. The generation of chemotactic factor can be prevented by heating of the serum, addition of EDTA or yeast phosphomannan. The generation of the chemotactic activity is explained by the fixation of piglet complement system which is activated by rough bacteria even in the absence of detectable antibodies in newborn sera.     

Distribution of ABCB1, CYP3A5, CYP2C19, and P2RY12 gene polymorphisms in a Mexican Mestizos population

The aim of the present study was to establish the gene frequency of six polymorphisms of the ABCB1, CYP3A5, CYP2C19, and P2RY12 genes in a population resident of Mexico City. The proteins encoded by these genes have been associated with the absorption, and biotransformation of clopidogrel. The ABCB1 T3435C, CYP3A5 V3* A6986G, P2RY12 G52T, P2RY12 C34T, CYP2C19 V2* and V3* (positions G681A and G636A, respectively), polymorphisms were analyzed by 5′ exonuclease TaqMan genotyping assays in a group of 269 healthy unrelated Mexican Mestizo individuals. The CYP2C19 V3* G636A polymorphism was not detected in the Mexican Mestizos population. However, the studied population presented significant differences (P < 0.05) in the distribution of the T3435C, A6986G, G681A, G52T and C34T polymorphisms when compared to reported frequencies of Amerindian of South America, Caucasian, Asian, and African populations. In summary, the distribution of the ABCB1, CYP3A5, CYP2C19, and P2RY12 gene polymorphisms distinguishes to the Mexican Mestizos population from other ethnic groups.     

Neutralization of bacterial viruses by antibodies of young animals

The cofactor activity of complement of precolostral newborn pig serum was compared with the haemolytic and bactericidal activity. Using current decomplementation treatment it was found that all three types of activity are destroyed in parallel i.e. haemolytic, bactericidal and cofactor (heat inactivation, EDTA, ammonia, trypsin, adsorption to zymosan or immune precipitate). In an ultracentrifugation analysis the haemolytic, bactericidal and cofactor activities showed the same sedimentation pattern. Using gel filtration on a Sephadex G 200 column, the cofactor activity was found in the fraction which contained all basic complement components. On the basis of these results it can be concluded that the complement of piglet serum acts as a factor which potentiates the neutralization of bacteriophage by the sera of the newborn.     

Immunological studies of the anticryptococcal factor of normal human serum

Preliminary studies have shown a very high inhibitory activity in the alpha₂ and gamma zone of human serum towards the growth of Cryptococcus neoformans. These findings are now corroborated by single radial immunodiffusion tests, which showed the some loss of IgA and IgM globulins and of the other three globulin fractions (ceruloplasmin, alpha₂ macroglobulin and alpha₂ HS glycoprotein) which migrate in the alpha₂ zone. The data was obtained by single radial immunodiffusion tests. The losses were not statistically significant however. No change in the immunoglobulin content of the sera kept for 6 days in contact with a heat-killed suspension of C. neoformans was noted. These findings suggest, that the inhibitory activity of the normal human serum on the in-vitro growth of C. neoformans is due to the above mentioned globulin fractions and not to a single specific factor.     

Elatoside C protects against hypoxia/reoxygenation-induced apoptosis in H9c2 cardiomyocytes through the reduction of endoplasmic reticulum stress partially depending on STAT3 activation

Endoplasmic reticulum (ER) stress-induced apoptosis has been suggested to contribute to myocardial ischemia–reperfusion (I/R) injury. Elatoside C is one of the major triterpenoid compounds isolated from Aralia elata that is known to be cardioprotective. However, its effects on I/R injury to cardiac myocytes have not been clarified. This study aimed to investigate the possible protective effect of Elatoside C against hypoxia/reoxygenation (H/R)-induced H9c2 cardiomyocyte injury and its underlying mechanisms. H9c2 cardiomyocytes were subjected to H/R in the presence of Elatoside C. Our results showed that Elatoside C (25 μM) treatment provided significant protection against H/R-induced cell death, as evidenced by improved cell viability, maintained mitochondrial membrane potential, diminished mitochondrial ROS, and reduced apoptotic cardiomyocytes (P < 0.05). These changes were associated with the inhibition of ER stress-associated apoptosis markers (GRP78, CHOP, Caspase-12 and JNK), as well as the increased phosphorylation of STAT3 and an increased Bcl2/Bax ratio. Moreover, these effects of Elatoside C were prevented by the STAT3 inhibitor Stattic. Taken together, these results suggested that Elatoside C can alleviate H/R-induced cardiomyocyte apoptosis most likely by activating the STAT3 pathways and reducing ER stress-associated apoptosis.     

Functional identification of ELO-like genes involved in very long chain fatty acid synthesis in Arabidopsis thaliana

Very long chain fatty acids (VLCFAs) are essential lipid components in many plants. 3-Ketoacyl-CoA synthase (KCS) catalyzes the condensation reaction to form 3-ketoacyl-CoA in VLCFA synthesis. AtELO4 has been reported to be involved in VLCFA synthesis, functioning as a KCS in Arabidopsis. However, no studies on other three AtELO members have been reported. Here, we initially found by real-time PCR in Arabidopsis thaliana (L.) Heynh. that AtELO1, AtELO3, and AtELO4 displayed characteristic expression patterns, but AtELO2 was nearly expressed in any organ. Then the transient expression of ELO-like-eGFP fusions in Arabidopsis green leaf protoplasts showed that AtELO1, AtELO3, and AtELO4 were localized in the endoplasmic reticulum (ER), where VLCFA synthesis took place. Finally, we found that the contents of all fatty acids were decreased by 10–20% in seeds of atelo1 T-DNA insertion mutants. In seeds of Pro35S:AtELO1 plants, the levels of all remaining components, except C20:0 and C20:3, were significantly increased. Taken together, our study revealed biological functions of AtELO members and might lay the foundation for further genetic manipulations to generate oil crops with the high oil content.     

The pathogenicity of an entomopoxvirus fromOthnonius batesi [Col.: Scarabaeidae] and its possible use as a control agent

In a natural population ofOthnonius batesi Oll. at Glen Innes, N.S.W., 4.0% of larvae in 1972 and 2.2% in 1973 were exhibiting symptoms of virus infection, whilst 0.8% of pupae and adults from the same population were infected in 1972. These figures, and field observations of infected larvae, suggested that the pathogenicity of the virus was low. In laboratory experiments withO. batesi the infection had little effect on mortality, and no significant effect on duration of the first instar, food intake, or larval growth. The vast accumulation of virus in the fat body probably results in mortality prior to, or during, the pupal period. Temperature had a marked effect on both % infection (optimum 30°C) and production of viral spheroids (optimum 25°C). Very little viral development occurred below 20°C. In a host range study onlyO. batesi, Rhopaea verreauxi Blanch. andR. morbillosa Blkb. were infectedper os. The possible use of the virus as a control agent is discussed.     

Conditional rifampicin sensitivity of arif mutant ofEscherichia coli: rifampicin induced changes in transcription specificity

Arif mutantof Escherichia coli that exhibits medium and temperature-dependent sensitivity to rifampicin is described. In the absence of rifampicin, this strain grows in minimal and rich media at 30°C and 42°C. In its presence it is viable in rich medium at both temperatures, but in minimal medium only at 30°C. In minimal-rifampicin medium at the higher temperature, RNA synthesis is decreased. The addition of certain divalent salts (MgSO₄, CaCl₂, BaCl₂) in excess, or chelators (EDTA, EGTA, o-phenanthrolein) greatly increase viability in minimal-rifampicin medium at 42°C. Excess MgSO₄ (10 mM) also increases the rate of RNA synthesis in the same medium. A model is proposed wherein therif mutation is suggested to cause a structural change in RNA polymerase that allows the binding of rifampicin and other ligands at 42°C. Rifampicin-binding is suggested to alter the conformation of RNA polymerase, impairing its ability to express genes required for growth in minimal medium. Implicit in this view is the assumption that these genes are structurally different from those expressed in rich medium in respect of certain template features recognized by RNA polymerase.     

Counterimmunoelectrophoresis as a routine mycoserological procedure

Counterimmunoelectrophoresis (CIE) has been compared in a diagnostic laboratory with agar gel double diffusion (DD) as a routine procedure for detection of antibodies to pathogenic and allergenic fungi and actinomycetes. It was shown to be of particular value in detecting antibodies to Aspergillus fumigatus. Thus 72 of 106 sera in which precipitins were detected were positive by CIE alone. Some sera were positive only by CIE to antigens prepared from Histoplasma capsulatum, Allescheria boydii, Candida albicans and C. parapsilosis.     

Response of the population size and community structure of Paenibacillus spp. to different fertilization regimes in a long-term experiment of red soil

Background and aims

Paenibacillus spp. are widely considered to impact the fertility and health of soil. The aim of this study was to evaluate how different fertilization regimes affect the population size and community structure of Paenibacillus spp. over a long period of time in red soil.

Methods

Soil samples were collected from a long-term experiment and were then analyzed using real-time PCR and PCR-DGGE. The correlation analysis, PCA and RDA were used to explore the relationships among Paenibacillus spp. population, community structure and soil properties in different treatments.

Results

The pH was seriously decreased only by the application of chemical fertilizer. The largest population of Paenibacillus spp. was found in the soil treated with organic fertilizer application, while the richest diversity was observed in the soil treated only with the chemical fertilizer. The Paenibacillus spp., Paenibacillus alkaliterrae, Paenibacillus campinasensis, and Paenibacillus xylanilyticus were found in all treatments. Paenibacillus castaneae was found in the soil treated with NPK, and Paenibacillus pabuli was specifically observed in the lime-amended treatment. Paenibacillus taichungensis and Paenibacillus prosopidis were detected in the soil treated with only chemical fertilizer. Except for the ammonium and pH, all the tested soil fertility parameters (total C, total N, nitrate, available K and available P) could significantly affect both the Paenibacillus spp. population number and diversity. The soil pH was significantly correlated with Paenibacillus spp. diversity only.

Conclusions

Our results indicate that the different long-term fertilization regimes have varied impact on both the Paenibacillus spp. population size and the diversity of the community associated with the soil properties tested. These results can help to enrich the information on the response of beneficial soil microbes to different long-term fertilization regimes.     

Expression profiles and association analysis with growth traits of the MyoG and Myf5 genes in the Jinghai yellow chicken

In order to analyze the association of single nucleotide polymorphisms (SNPs) in the MyoG and Myf5 genes with chicken growth traits, PCR-SSCP approach was used to detect the (SNPs). The general linear model was used to analyze gene interaction and genetic effects between different genotypes and growth traits of the Jinghai yellow chicken. For the MyoG gene, three genotypes (AA, AB and BB) were detected in the Jinghai yellow chicken population. Gene sequencing revealed one mutation (T36C) in the genotype BB in comparison to the genotype AA. For the Myf5 gene, three genotypes (CC, CD and DD) were detected in the Jinghai yellow chicken population. Gene sequencing revealed one mutation (A1313G) in the genotype DD in comparison to the genotype CC. Gene interaction effect has significant influence on 6, 8-week-weight and 300-day-weight. The least square analysis showed that individuals with BB genotype of the MyoG gene had higher bodyweight at 2, 4, 10, 12, 14 and 16 weeks compared to individuals with AA and AB genotypes. Individuals with CD genotype of the Myf5 gene had higher birth weight than individuals with CC genotype (P < 0.05). The interactive genotype AB*DD performs well at 6, 8 weeks and 300 days bodyweight. The results suggested that SNPs of the MyoG and Myf5 genes had certain effects on growth traits of the Jinghai yellow chicken.     

Genetic variability of complement receptor on human erythrocytes

Erythrocytes from healthy men were examined for the presence of complement (C3b) receptor using haemagglutination assay with human aggregated IgG (aggIgG) and guinea pig complement. The results were expressed as the intensity of haemagglutination that corresponded to the C3b receptor sites density as evidenced by radioimmunobinding results. Among normal men three phenotypes of complement receptor (CR) were distinguished: high (CR^h/CR^h) phenotype corresponding to strong agglutination, an intermediate (CR^h/CR^I) producing weak agglutination and low phenotype (CR^I/CR^I) that gave no agglutination. In a population of 517 normal men these three phenotypes occurred in 63.8, 30.6 and 5.6%, respectively. Frequencies of the genes responsible for high (CR^h) and low (CR^I) expression of erythrocyte C3b receptor were 0.791 and 0.209, respectively.     

Immune response of mice to the Thy-1.1 antigen: Effect of theIr-5 alleles studied in 129/J and B10.129 (6M) mice

The primary immune response to the Thy-1.1 antigen was measured by a plaque assay that detected cells producing antibodies lytic for AKR thymocytes. B10.129(6M) mice carrying theH-2 complex of an intermediate responder (129) on a low-responder (B10) background, were low responders. Studies employing different F₁ hybrids and segregating generations of 129/J and 6M mice indicated that differences in responsiveness of those two strains depend on alleles at a single locus, loosely linked to theH-2 complex. These results lend further support to the previously advanced concept that the expression of theIr-Thy-1 allcles controlling the response to the Thy-1.1 antigen is influenced by the alleles at theIr-5 locus. In addition, studies employing F₁ hybrids produced through matings of 129/J, 6M, C3H.B10 and C57BL/6J mice to a panel of inbred strains suggested that in regard to the responsiveness to the Thy-1.1 antigen, 129/J and 6M mice are phenotypically, and presumably genotypically, similar to C3H.B10 and C57BL/6J mice, respectively.     

Production of the mosquito-parasitic fungus,Coelomomyces dodgei,through synchronized infection and growth of the intermediate copepod host,Cyclops vernalis

High yields of the copepodCyclops vernalis infected with the mosquito-parasitic fungusCoelomomyces dodgei Couch & Dodge were obtained by infecting nauplii in large synchronously developing populations. Exposure of 2000 48 or 72 h old nauplii to 6×10³ sporangia at the time of meiospore release yielded ca. 1500 infected copepods. Based on yields of infected copepods, susceptibility ofC. vernalis toC. dodgei decreased as copepods developed. Infection rates were 75% for copepods exposed as 48 or 72 h old nauplii but declined to 32 and 9.6%, respectively, for those exposed as copepodids or adults. The relevance of these results for domestication of other species ofCoelomomyces and studies on non-target organisms is discussed, and improved procedures for routine production ofC. dodgei are described.     

NaturalMycoplasma arthritidis infection in mice

B6C3F₁ mice from a hybrid production colony frequently were serologically positive by enzyme-linked immunosorbent assay (ELISA) and consistently negative by culture forMycoplasma pulmonis. Subsequently, 162 mice were obtained and intensively studied using an expanded group of cultural procedures, ELISA, and histopathology. Lesions attributable to mycoplasma infection were not found, butMycoplasma arthritidis was isolated from 20 mice. TheM. pulmonis ELISA was positive (IgM, IgG, or both) in 113 mice. Selected sera were tested simultaneously in both theM. pulmonis ELISA and in an ELISA usingM. arthritidis antigen, and were found to be positive in both the IgM and IgG classes in both ELISAs. Thus, cross-reacting antibody was produced in mice naturally infected withM. arthritidis, confirming previous observations based on experimental infections. To our knowledge, this is the first report of naturalM. arthritidis infection in laboratory mice.     

Interaction between smoking and functional polymorphism in the TGFB1 gene is associated with ischaemic heart disease and myocardial infarction in patients with rheumatoid arthritis: a cross-sectional study

Introduction

Transforming growth factor-beta1 (TGF-beta1) is a pleiotropic cytokine that plays important roles in immunity and inflammation. Some studies have suggested that polymorphism in the TGFB1 gene is associated with heart disease in the general population. The purpose of the present study was to determine whether common single-nucleotide polymorphisms (SNP) in the TGFB1 gene are associated with ischaemic heart disease (IHD) and/or myocardial infarction (MI) in patients with rheumatoid arthritis (RA), and to investigate the influence of smoking on any association.

Methods

PCR-based assays were used to determine the genotypes of TGFB1 SNPs including TGFB1-509 C/T (rs1800469, in the promoter region), +868 T/C (rs1800470, in exon 1) and +913 G/C (rs1800471, in exon 1) in 414 subjects with established RA. Genotyping for the +868 SNP was also carried out on a second study population of RA patients (n = 259) with early disease. Serum levels of TGF-beta1 were measured using a commercial ELISA kit. Smoking history and IHD/MI status were obtained on each patient. Associations with IHD/MI were assessed using contingency tables and logistic regression analyses.

Results

The heterozygous genotype of TGFB+868 was associated with an increased risk of IHD (OR 2.14, 95% CI 1.30 - 3.55) and MI (OR 2.42, 95% CI 1.30-4.50), compared to the homozygous genotypes combined. Smoking was an independent risk for IHD and MI, and evidence of interaction between smoking and TGFB+868 was found. Multivariate analyses indicated that the strongest associations with IHD and MI were due to the combined effect of the TGFB1+868 TC genotype and smoking (OR 2.75, 95% CI 1.59-4.75; and OR 2.58 95% CI 1.33-4.99, respectively), independent of other cardiovascular risk factors. The association of the +868 TC genotype and evidence of +868 TC-smoking interaction with IHD were replicated in a second population of RA patients with early disease. Serum TGF-beta1 levels were not associated with TGFB1 genetic variations, smoking or IHD/MI status.

Conclusions

Interaction between smoking and polymorphism in the TGFB1 gene may influence the risk of IHD and MI in patients with RA.