SUMMARY Rabbit polyclonal hyperimmune antibodies to Yersinia pestis , and a mouse monoclonal antibody against the capsular antigen fraction 1 (F1) were compared in immunofluorescence (IF) tests. Fluorescent antibody conjugates were prepared from polyclonal antisera to four F1 positive Y. pestis strains; the conjugated antibody to strain A1122 gave the strongest IF staining of F1 positive and F1 negative Y. pestis strains. Indirect assays were rejected in favour of direct assays utilizing polyclonal and monoclonal reagents because the increased background staining reduced the effective contrast of bacterial visualisation. Polyclonal conjugates gave fairly homogeneous staining of Y. pestis bacterial populations, but in monoclonal assays a skew distribution of fluorescence intensity was observed, the majority of bacteria being poorly stained. The proportion of cells stained well by the monoclonal sufficed for easy identification of Y. pestis of the F1 positive phenotype however, and staining was not affected by washing the bacteria or treating them with formaldehyde. Y. pestis strains of the F1 positive genotype reacted with the monoclonal if bacteria were grown at 37 °C but not if the growth temperature was reduced to 25°C thus preventing capsule production. The polyclonal conjugate reacted with bacteria of these strains that had been grown at either temperature. Strains of F1 negative genotype grown at either temperature. Strains of F1 negative genotype grown at either temperature reacted with the polyclonal conjugate but not with the monoclonal. Cross reactions between the polyclonal reagents and Y. enterocolitica biovar 2, serovar O 8 could not be removed by selective absorption; however, the monoclonal antibody gave no cross reaction. The F1 phenotypic status of bacterial preparations was verified by ELISA measurement of the fraction 1 antigen concentration. Antigen levels for F1 positive and F1 negative phenotypes differed by about three logs for suspensions of Y. pestis harvested from solid media. The polyclonal and monoclonal direct IF tests applied to spleen and blood smears of laboratory mice infected with Y. pestis were able to differentiate between lethal infection with an F1 positive strain carrying all four classical virulence determinants, an F1 positive vaccine strain, and an F1 negative strain.
Forestry best management practices (BMPs) are used to reduce sedimentation from forest stream crossings. Three BMP treatments (BMP−, BMP-std, and BMP+) were applied to three forest road stream crossings (bridge, culvert, and ford). BMP− did not meet existing BMP guidelines, BMP-std met standard recommendations, and BMP+ treatments exceeded recommendations. Following BMP applications, three simulated rainfall intensities (low, medium, and high) were applied in order to evaluate sediment delivery from crossing type and BMP level. During rainfall simulation, sediment concentrations (mg/L) were collected with automated samplers and discharge (L/s) was estimated to calculate total sediment loading. Costs of stream crossings and BMP levels were also quantified. Mean sediment associated with the three stream crossings were 3.38, 1.87, and 0.64 Mg for the BMP−, BMP-std, and BMP+ levels, respectively. Ford, culvert, and bridge crossings produced 13.04, 12.95, and 0.17 Mg of sediment during construction, respectively. BMP enhancement was more critical for sediment control at the culvert and ford crossings than at the bridge. Respective costs for BMP−, BMP-std, and BMP+ levels were $5,368, $5,658, and $5,858 for the bridge; $3,568, $4,166 and $4,595 for the culvert; and $180, $420 and $1,903 for the ford. Costs and sediment values suggest that current standard BMP levels effectively reduce stream sediment while minimizing costs.
A fragment of DNA containing the gene coding for the phospholipase C (alpha-toxin) of Clostridium perfringens was cloned into Escherichia coli. The cloned DNA appeared to code only for the alpha-toxin and contained both the coding region and its associated gene promoter. The nucleotide sequence of the cloned DNA was determined, and an open reading frame was identified which encoded a protein with a molecular weight of 42,528. By comparison of the gene sequence with the N-terminal amino acid sequence of the protein, a 28-amino-acid signal sequence was identified. The gene promoter showed considerable homology with the E. coli sigma 55 consensus promoter sequences, and this may explain why the gene was expressed by E. coli. The cloned gene product appeared to be virtually identical to the native protein. A 77-amino-acid stretch that was close to the N terminus of the alpha-toxin showed considerable homology with similarly located regions of the Bacillus cereus phosphatidylcholine, preferring phospholipase C and weaker homology with the phospholipase C from Pseudomonas aeruginosa.
Recent decisions by the United States Supreme Court and United States Environmental Protection Agency (EPA) have re-emphasized the importance of forestry best management practices (BMPs) at stream crossings. Stream crossings are potential major sources of sediment due to their direct connectivity between the potential erosion source and the stream, which eliminates potential sediment reduction provided by filter/buffer strips and stream side management zones. The effectiveness of stream crossing BMPs for sediment control were tested for a permanent bridge crossing, culvert crossing, and improved ford crossing on three first-order streams in the Virginia Piedmont using rainfall simulation. The three crossings were located on a low standard legacy road having unimproved ford crossings before experimentation. All legacy fords received three levels of rainfall intensity via simulation prior to crossing installation. Following crossing installation, rainfall simulations were performed at each of the crossings under the following three treatments: (1) minimal levels of BMP erosion control (Low); followed by (2) installation of BMPs recommended by the Virginia BMP Manual (Medium); and (3) erosion control measures beyond the Virginia BMP Manual (High). Stream sediment (TSS) was monitored upstream and downstream during rainfall simulations to determine total sediment contribution from each individual crossing. The comparison of minimal BMPs, recommended BMPs, and extensive protection provides insight into the erosion associated with the crossing types and the effectiveness of current BMPs for nonpoint source pollution (NPSP) reduction. The Culvert crossing produced a sediment concentration (2.9 g/L) that was double the concentration produced by the Ford crossing (1.4 g/L) and over 10 times the concentration of the Bridge crossing (0.2 g/L).
Abstract : Introduction of the F-lac episome from Escherichia coli into Pasteurella pseudotuberculosis yielded a strain of P. pseudotuberculosis F-lac that could transfer its genes by conjugation to Pasteurella pestis. The selected P. pseudotuberculosis marker arginine+ was transferred at frequencies between 1/ 1,000,000 and 1/10,000,000 per donor cell. Unselected donor characters such as melibiose of rhamnose fermentation, phage sensitivity, and ureas production appeared at different frequencies in the recombinants. The transferred markers were unrelated to the transfer of F-lac or its subsequent loss from the recipient cell.
As a result of the extensive number of applications of silver nanoparticles (AgNPs), their potential impacts, once released into the environment, are of concern. The toxicity of AgNPs was reported to be dependent on various factors such as particle size, shape and capping agent. Although these factors may play a role in AgNPs toxicity, the results presented herein suggest that surface charge is one of the most important factors that govern the toxicity of AgNPs. In the current study, the toxicity of four AgNPs representing various surface charging scenarios ranging from highly negative to highly positive was investigated. These AgNPs were (1) uncoated H(2)-AgNPs, (2) citrate coated AgNPs (Citrate-AgNPs), (3) polyvinylpyrrolidone coated AgNPs (PVP-AgNPs), and (4) branched polyethyleneimine coated AgNPs (BPEI-AgNPs). Our results clearly demonstrate that the AgNPs exhibited surface charge-dependent toxicity on the bacillus species investigated. Furthermore, ultrafiltration membranes were utilized to purify the AgNPs suspensions from residual impurities prior to the introduction to the microbes. This step was crucial in determining the true AgNPs toxicity and is either missing or not explicitly mentioned in most of the reported toxicity studies.
SUMMARY: Pasteurella pseudotuberculosis strain 321V accepted the F'lac episome from Escherichia coli strain 23.10S and behaved as a gene donor in crosses with several different auxotrophs of P. pseudotuberculosis. Some selected donor markers were transferred at frequencies of 10−4-10−5 per donor cell while others appeared not to be transferred. Up to 40% of recombinants were Lac+. Selected recombinants showed differing unselected marker frequencies with differing selected markers; those obtained by using double marker selection showed increased unselected marker frequencies. Some alternative explanations for the origin of recombinants (syntrophic growth, mixed clones, multiple recipient reversions) were not supported by experiment.
'%3e%3cpath fill='%23012169' d='M0 0v30h60V0z'/%3e%3cpath stroke='%23fff' stroke-width='6' d='m0 0 60 30m0-30L0 30'/%3e%3cpath stroke='%23C8102E' stroke-width='4' d='m0 0 60 30m0-30L0 30' clip-path='url(%23b)'/%3e%3cpath stroke='%23fff' stroke-width='10' d='M30 0v30M0 15h60'/%3e%3cpath stroke='%23C8102E' stroke-width='6' d='M30 0v30M0 15h60'/%3e%3c/g%3e%3c/svg%3e)
