Genetic basis for colonial variation in Neisseria gonorrhoeae.

When the piliated colony types of Neisseria gonorrhoeae, which predominate in recent isolates, were nonselectively subcultured in vitro, they gave rise to large numbers of nonpiliated, avirulent colonial variants. Evidence is presented to show that most of this variation occurs after active growth has ceased and that the variation is sensitive to the action of deoxyribonuclease. We suggest that this variation is a result of transformation. A second variation in colonial morphology involved differing levels of "colony opacity-associated proteins" in the outer membrane. This variation was also inhibited by the presence of deoxyribonuclease, but the genetic basis for it is not as yet clear.     

In vivo regulation of chromosomal beta-lactamase in Escherichia coli.

Chromosomal beta-lactamase, a periplasmic enzyme of Escherichia coli, was studied with respect to its regulation in vivo. Both the activity and the amount of beta-lactamase increased with growth rate. During a nutritional shift-down, chromosomal beta-lactamase activity followed stable ribonucleic acid accumulation. After a nutritional shift-up the differential rate of beta-lactamase synthesis did not increase immediately (like stable ribonucleic acid), but did increase after a lag period of 30 min. To determine whether beta-lactamase was under stringent control, strains carrying a temperature-sensitive valyl-transfer ribonucleic acid synthetase and differing only in the allelic state of the relA gene were shifted from a permissive to a semipermissive temperature. No influence by the relA gene product was found on beta-lactamase synthesis. The regulation of this periplasmic enzyme is discussed in relation to that of some components of the translational apparatus.     

Growth pattern and cell division in Neisseria gonorrhoeae.

The gram-negative coccus Neisseria gonorrhoeae was found to grow regularly in at least two dimensions. Growth proceeded at a linear rate sequentially in each dimension. Growth in the second dimension (former width) was initiated slightly before the pole-division plane distance equalled the cell width. Penicillin treatment localized presumptive growth zones to the existing septum region. It was suggested that new growth zones were always formed perpendicular to the longitudinal axis created in the incipient daughter cells of a dividing coccus. Neither penicillin nor nalidixic acid induced filaments of N. gonorrhoeae. Such structures could nevertheless be formed in the rod-shaped species Neisseria elongata. N. gonorrhoeae divides by septation; however, complete septal structures with separated cytoplasms were rather infrequent. It is proposed that N. gonorrhoeae be regarded as a short rod which always extends parallel to the actual longitudinal axis and which never undergoes a rod-sphere-rod transition.     

Estimation of heterozygosity for single-probe multilocus DNA fingerprints

In spite of the increasing application of DNA fingerprinting to natural populations and to the genetic identification of humans, explicit methods for estimation of basic population genetic parameters from DNA fingerprinting data have not been developed. Contributing to this omission is the inability to determine, for multilocus fingerprinting probes, relatively important genetic information, such as the number of loci, the number of alleles, and the distribution of these alleles into specific loci. One of the most useful genetic parameters that could be derived from such data would be the average heterozygosity, which has traditionally been employed to measure the level of genetic variation within populations and to compare genetic variation among different loci. We derive here explicit formulas for both the estimation of average heterozygosity at multiple hypervariable loci and a maximum value for this estimate. These estimates are based upon the DNA restriction-pattern matrices that are typical for fingerprinting studies of humans and natural populations. For several empirical data sets from our laboratory, estimates of average and maximal heterozygosity are shown to be relatively close to each other. Furthermore, variances of these statistics based on simulation studies are relatively small. These observations, as well as consideration of the effect of missing alleles and alternate numbers of loci, suggest that the average heterozygosity can be accurately estimated using phenotypic DNA fingerprint patterns, because this parameter is relatively insensitive to the lack of certain genetic information.      

Maladjusted Host Immune Responses Induce Experimental Cerebral Malaria-Like Pathology in a Murine Borrelia and Plasmodium Co-Infection Model

In the Plasmodium infected host, a balance between pro- and anti-inflammatory responses is required to clear the parasites without inducing major host pathology. Clinical reports suggest that bacterial infection in conjunction with malaria aggravates disease and raises both mortality and morbidity in these patients. In this study, we investigated the immune responses in BALB/c mice, co-infected with Plasmodium berghei NK65 parasites and the relapsing fever bacterium Borrelia duttonii. In contrast to single infections, we identified in the co-infected mice a reduction of L-Arginine levels in the serum. It indicated diminished bioavailability of NO, which argued for a dysfunctional endothelium. Consistent with this, we observed increased sequestration of CD8+ cells in the brain as well over expression of ICAM-1 and VCAM by brain endothelial cells. Co-infected mice further showed an increased inflammatory response through IL-1β and TNF-α, as well as inability to down regulate the same through IL-10. In addition we found loss of synchronicity of pro- and anti-inflammatory signals seen in dendritic cells and macrophages, as well as increased numbers of regulatory T-cells. Our study shows that a situation mimicking experimental cerebral malaria (ECM) is induced in co-infected mice due to loss of timing and control over regulatory mechanisms in antigen presenting cells.     

Influenza A virus‐mediated priming enhances cytokine secretion by human dendritic cells infected with Streptococcus pneumoniae

Secondary infections with Streptococcus pneumoniae (SP) are frequently observed following influenza A virus (IAV) infection and have a substantial impact on global health. Despite this, the basis for the disease progression is incompletely understood. To investigate the effect of co‐infection on human monocyte‐derived dendritic cells (MDDCs) we analysed the expression of clinically important pro‐inflammatory and immune‐modulatory cytokines. IAV infection or treatment with supernatants from IAV‐infected cell cultures resulted in priming of the DCs which subsequently influenced the production of IL‐12p70, as well as IL‐6, following SP infection. Co‐infection of the same cell was not required but this effect was dependent on the time, dose and duration of the infections, as well as pathogen viability, bacterial uptake and endosome acidification. Bacterially infected cells were characterized as the main producers of IL‐12p70. Finally, we showed that type I interferons were primarily responsible for the priming of IL‐12p70 that was observed by infection with IAV. These results provide a probable mechanism for the elevated levels of particular cytokines observed in IAV and SP co‐infected cell cultures with implications for the pathogenic outcome observed during in vivo infection.     

Contribution of subsurface peat to CO2 and CH4 fluxes in a neotropical peatland

Tropical peatlands play an important role in the global carbon cycling but little is known about factors regulating carbon dioxide (CO₂) and methane (CH₄) fluxes from these ecosystems. Here, we test the hypotheses that (i) CO₂ and CH₄ are produced mainly from surface peat and (ii) that the contribution of subsurface peat to net C emissions is governed by substrate availability. To achieve this, in situ and ex situ CO₂ and CH₄ fluxes were determined throughout the peat profiles under three vegetation types along a nutrient gradient in a tropical ombrotrophic peatland in Panama. The peat was also characterized with respect to its organic composition using ¹³C solid state cross‐polarization magic‐angle spinning nuclear magnetic resonance spectroscopy. Deep peat contributed substantially to CO₂ effluxes both with respect to actual in situ and potential ex situ fluxes. CH₄ was produced throughout the peat profile with distinct subsurface peaks, but net emission was limited by oxidation in the surface layers. CO₂ and CH₄ production were strongly substrate‐limited and a large proportion of the variance in their production (30% and 63%, respectively) was related to the quantity of carbohydrates in the peat. Furthermore, CO₂ and CH₄ production differed between vegetation types, suggesting that the quality of plant‐derived carbon inputs is an important driver of trace gas production throughout the peat profile. We conclude that the production of both CO₂ and CH₄ from subsurface peat is a substantial component of the net efflux of these gases, but that gas production through the peat profile is regulated in part by the degree of decomposition of the peat.     

Evaluation of approaches for measuring taxonomic completeness of lake profundal macroinvertebrate assemblages

1. Benthic macroinvertebrates (MI) are commonly used to assess freshwater ecosystems with the reference condition approach. Such assessments necessitate control for natural community variation, either by categorical typologies or by predictive models that have been widely and successfully developed for running water biota but not previously for lake profundal invertebrates. 2. We evaluated four modelling techniques [multivariate regression tree (MRT), limiting environmental differences, nonparametric multiplicative regression (NPMR) and River Invertebrate Prediction And Classification System (RIVPACS) and the operative Finnish lake typology for assessing taxonomic completeness (observed‐to‐expected number of taxa, O/E) of profundal MI assemblages. We used data from 74 and 33 minimally disturbed reference lake basins for calibration and validation of the approaches, respectively, and 72 test basins subject to various anthropogenic pressures to evaluate sensitivity to detect impact. Either all predicted taxa (threshold probability of capture P_t = 0+) or only those predicted to be captured with ≥0.25 probability were used to calculate O/E. 3. With P_t = 0.25, all four modelling approaches were accurate (mean O/E = 0.966–1.053) but imprecise (SD of O/E = 0.279–0.304) in predicting the fauna actually observed in validation sites. All models were subtly more precise than a null model (mean 1.038, SD 0.343) or the typology (1.046, 0.327). The taxon‐specific NPMR model was slightly more precise than the other three models based on site groupings. 4. The O/E values correlated relatively weakly (r = 0.55–0.86) among the approaches, which thus produced contrasting lake‐specific assessments, despite their seemingly comparable performances. Indeed, typology, suggesting that MI assemblages were impaired in 56% of test sites, was more sensitive than the other approaches (26–46%) as an indicator of human‐induced deterioration. However, this greater ostensible sensitivity seemed to be biased, as lake morphometry, a main driver of natural community variation, remained uncontrolled by the typology. 5. Generally, our exercise illustrates the inconclusiveness of the common validation criteria for the assessment methods. The apparent poor predictability of the profundal fauna, irrespective of the method, may partly stem from large observation error, which could be alleviated by more intensive sampling. However, instead of an O/E‐taxa index, some other metric encompassing quantitative aspects might be preferable for assessing these species‐poor communities.