Dynamic change in natural killer cell type in the human ocular mucosa in situ as means of immune evasion by adenovirus infection.

The most severe form of virus-induced inflammation at the ocular surface is epidemic keratoconjunctivitis (EKC), often caused by group D human adenoviruses (HAdVs). We investigated the dynamics and mechanisms of changes in natural killer (NK) cell types in the human ocular mucosal surface in situ over the course of infection. In the acute phase of infection, the mature CD56(dim)NK cells that comprise a major subpopulation in the normal human conjunctiva are replaced by CD56(bright)NK cells recruited to the ocular surface by chemokines produced by the infected epithelium, and NKG2A-expressing CD56(dim) and CD56(bright) NK cells become the major subpopulations in severe inflammation. These NK cells attracted to the mucosal surface are however incapable of mounting a strong antiviral response because of upregulation of the inhibitory ligand human leukocyte antigen-E (HLA-E) on infected epithelium. Furthermore, group D HAdVs downregulate ligands for activating NK cell receptors, thus rendering even the mature NKG2A(-)NK cells unresponsive, an immune-escape mechanism distinct from other adenoviruses. Our findings imply that the EKC-causing group D HAdVs utilize these multiple pathways to inhibit antiviral NK cell responses in the initial stages of the infection.

Screening for childhood obesity: international vs population-specific definitions. Which is more appropriate?

AIM: The objectives of this study are: (1) to study the relation between body mass index (BMI), percentage-weight-for-height (PWH) and percentage body fat (PBF) in Singaporean Chinese children; (2) to assess the applicability of an international definition of obesity (the International Obesity Task Force (IOTF) BMI) as a screening tool to detect childhood obesity, as compared with the current Singapore population-specific definition using PWH. METHODS: A total of 623 Chinese children aged 6-11 y (321 males, 302 females) were recruited from a school by proportionate (40%) stratified random sampling. BMI and PWH were calculated from weight and height, while PBF was derived using leg-to-leg bioelectrical impedance analysis. The strength of association among the three indices of obesity was assessed using Spearman's correlation coefficient. Obese children were defined as those above the 95th percentile of PBF in each age-gender-specific group. Sensitivity and specificity of IOTF-BMI cutoff values and PWH cutoff values were compared by testing their ability to correctly identify obese children. RESULTS: All three indices correlated well with one another (BMI:PWH r=0.83, BMI:PBF r=0.87, PWH:PBF r=0.76). Prevalence of obesity was lower using IOTF-BMI cutoffs (6.9%) than using PWH cutoffs (16.4%). The sensitivity and specificity of IOTF-BMI cutoff values were 75.0 and 96.0%, respectively, with sensitivity differing between boys (83.3%) and girls (66.6%) (P=0.35). In comparison, PWH cutoff values had higher sensitivity (91.6%) but lower specificity (86.6%), with no significant difference between the genders. CONCLUSION: IOTF-recommended BMI cutoff values had low sensitivity and may underestimate the local prevalence of childhood obesity. For screening purposes, we recommend that population-specific measures rather than international cutoff values be used.

Recruitment of a chromosomally encoded maleylacetate reductase for degradation of 2,4-dichlorophenoxyacetic acid by plasmid pJP4.

When Pseudomonas aeruginosa PAO1c or P. putida PPO200 or PPO300 carry plasmid pJP4, which encodes enzymes for the degradation of 2,4-dichlorophenoxyacetic acid (TFD) to 2-chloromaleylacetate, cells do not grow on TFD and UV-absorbing material with spectral characteristics of chloromaleylacetate accumulates in the culture medium. Using plasmid pRO1727, we cloned from the chromosome of a nonfluorescent pseudomonad, Pseudomonas sp. strain PKO1, 6- and 0.5-kilobase BamHI DNA fragments which contain the gene for maleylacetate reductase. When carrying either of the recombinant plasmids, pRO1944 or pRO1945, together with pJP4, cells of P. aeruginosa or P. putida were able to utilize TFD as a sole carbon source for growth. A novel polypeptide with an estimated molecular weight of 18,000 was detected in cell extracts of P. aeruginosa carrying either plasmid pRO1944 or plasmid pRO1945. Maleylacetate reductase activity was induced in cells of P. aeruginosa or P. putida carrying plasmid pRO1945, as well as in cells of Pseudomonas strain PKO1, when grown on L-tyrosine, suggesting that the tyrosine catabolic pathway might be the source from which maleylacetate reductase is recruited for the degradation of TFD in pJP4-bearing cells of Pseudomonas sp. strain PKO1.

The activation of mutagens in diesel particle extract with rat liver S9 enzymes.

Reductive activation of nitroaromatic compounds by bacterial enzymes contributes to the direct-acting mutagenicity of diesel particle extracts in the Salmonella mutation assay. In this study, the ability of mammalian enzymes to activate diesel particle extract was investigated with a rat liver S9 preparation using the nitroreductase-deficient strain TA98NR to detect active metabolic products. The use of diesel particle extract at concentrations greater than 50 micrograms per plate inhibited S9 enzyme activations in the mutation assay. In addition, the S9 preparation without NADPH present decreased the residual direct-acting mutagenicity of the particle extract. Activation by S9 enzymes of mutagens in diesel particle extract was evident as a difference in activity between assays with and without NADPH. 1-Nitropyrene, a nitroarene identified in diesel particle extracts, was also activated by NADPH-dependent S9 enzymes. S9 activation of both the particle extract and 1-nitropyrene was detected only when using much lower S9 concentrations than conventionally applied in the Salmonella/S9 assay. S9 enzyme activation of 1-nitropyrene is not merely a consequence of forming a mutagenic amine since 1-aminopyrene was less mutagenic than 1-nitropyrene in these assays. The NADPH-dependent activation of 1-nitropyrene is located in the microsomal fraction of the S9 preparation. However, activation of the diesel particle extract was more evident with the cytosol than with the microsomal fraction, demonstrating the involvement of yet other enzyme systems and extract components.

The role of nitroaromatic compounds in the direct-acting mutagenicity of diesel particle extracts.

The Salmonella mutation assay has been adapted to a number of experimental procedures used to characterize the chemical components and molecular mechanisms involved in the direct-acting mutagenicity of diesel particle extracts. The mutagens were found to be unreactive towards purified DNA. Mutagenic activity was decreased in nitroreductase-deficient bacteria and increased in the absence of oxygen, suggesting that bacterial enzymes active the mutagens. The extract was fractionated by thin layer chromatography using both normal and reverse phase silica gel plates. On normal silica plates, a separation was made between unsubstituted PAH, nitro-PAH and the more polar aromatic compounds. About a third of the mutagenic activity in the particle extract was recovered in fractions containing monosubstituted nitro-PAH compounds. The absorption spectra and reverse phase chromatographic properties indicate 1-nitropyrene is a predominant component but not the only mutagen in these fractions. The remaining activity was in the more polar fractions, and mutagenicity assays using anaerobic conditions and nitro-reductase-deficient bacteria suggest they contain other nitro-substituted compounds.

Survival of coxsackievirus B3 under diverse environmental conditions.

The survival of coxsackievirus B3 was studied under various conditions of incubation. The comparative study demonstrated that coxsackievirus B3 was stable for 24h (less than 0.4-log decrease in titer) when suspended at neutral pH (6 or 23 degrees C) in the presence of 0.25% bovine serum albumin in saline regardless of whether the preparations were subjected to evaporation. Bovine serum albumin provided increased stability to the virus for each of the conditions tested. At 37 degrees C, evaporation greatly reduced the virus infectivity between 6 and 20 h of incubation. Nevertheless, coxsackievirus B3 was found to be stable for at least 24 h under conditions similar to those of a household environment, and its presence represents a potential biohazard to nonimmune persons. These data provide a rationale for using coxsackievirus B3 as a model for investigating the role of environmental surfaces in the transmission of enteroviral diseases.

Characteristics of PRD1, a plasmid-dependent broad host range DNA bacteriophage.

Several distinctive properties of PRD1, an icosahedral plasmid-dependent phage, are described. The drug-resistance plasmid-dependent host range of PRD1 extends beyond the P incompatibility group and includes gram-negative bacteria containing plasmids of incompatibility groups N and W. PRD1 phage will infect pseudomonads and Enterobacteriaceae containing either a P or W incompatibility group plasmid. PRD1 adsorbs to the cell wall of R(+) bacteria and thus its infectivity indicates cell wall alterations by these drug-resistance plasmid groups. PRD1 nucleic acid is duplex DNA with an estimated molecular weight of 24 x 10(6). The appearance of PRD1 in electron micrographs is suggestive of lipid content in addition to its buoyant density of 1.348 in CsCl and its sensitivity to chloroform. The latent period of PRD1 varies with the R(+) host bacterial strain used for growth of the phage.