United States Bureau of Mines Bulletin 590

From Introduction: "The purpose of this Bulletin, in addition to presenting a history and description of Bureau activities in controlling these fires, is to provide the mining industry and the public with information concerning methods of controlling fires in inactive coal formation. The Bulletin is intended to serve as a guide to the public and more specifically to mining supervisory personnel and engineers who may be called upon to extinguish such fires.

United States Bureau of Mines Bulletin 596

From Introduction and Summary: "This bulletin presents the results obtained n-heptane by the petroleum thermodynamics laboratory of the Bartlesville, Okla., facility. Detailed results of five investigations are given; selected values if the low-temperature thermal properties are tabulated; and comparisons with results that were obtained in other laboratories are made. The findings of this extensive study will serve to characterize the low-temperature thermal data for over 100 compounds that will be presented in a later bulletin of this series.

Correction: Two stable variants of Burkholderia pseudomallei strain MSHR5848 express broadly divergent in vitro phenotypes associated with their virulence differences.

[This corrects the article DOI: 10.1371/journal.pone.0171363.]

Two stable variants of <i>Burkholderia pseudomallei</i> strain MSHR5848 express broadly divergent <i>in vitro</i> phenotypes associated with their virulence differences

<div><p><i>Burkholderia pseudomallei</i> (<i>Bp</i>), the agent of melioidosis, causes disease ranging from acute and rapidly fatal to protracted and chronic. <i>Bp</i> is highly infectious by aerosol, can cause severe disease with nonspecific symptoms, and is naturally resistant to multiple antibiotics. However, no vaccine exists. Unlike many <i>Bp</i> strains, which exhibit random variability in traits such as colony morphology, <i>Bp</i> strain MSHR5848 exhibited two distinct and relatively stable colony morphologies on sheep blood agar plates: a smooth, glossy, pale yellow colony and a flat, rough, white colony. Passage of the two variants, designated “Smooth” and “Rough”, under standard laboratory conditions produced cultures composed of > 99.9% of the single corresponding type; however, both could switch to the other type at different frequencies when incubated in certain nutritionally stringent or stressful growth conditions. These MSHR5848 derivatives were extensively characterized to identify variant-associated differences. Microscopic and colony morphology differences on six differential media were observed and only the Rough variant metabolized sugars in selective agar. Antimicrobial susceptibilities and lipopolysaccharide (LPS) features were characterized and phenotype microarray profiles revealed distinct metabolic and susceptibility disparities between the variants. Results using the phenotype microarray system narrowed the 1,920 substrates to a subset which differentiated the two variants. Smooth grew more rapidly <i>in vitro</i> than Rough, yet the latter exhibited a nearly 10-fold lower lethal dose for mice than Smooth. Finally, the Smooth variant was phagocytosed and replicated to a greater extent and was more cytotoxic than Rough in macrophages. In contrast, multiple locus sequence type (MLST) analysis, ribotyping, and whole genome sequence analysis demonstrated the variants’ genetic conservation; only a single consistent genetic difference between the two was identified for further study. These distinct differences shown by two variants of a <i>Bp</i> strain will be leveraged to better understand the mechanism of <i>Bp</i> phenotypic variability and to possibly identify <i>in vitro</i> markers of infection.</p></div

Western blot of LPS from <i>Bp</i> strains.

<p>Purified LPS from MSHR5848 Smooth and Rough, and <i>Bp</i> strain 1106a, were separated by SDS-PAGE and a western blot was done using monoclonal antibody (mAb) 11G3-1, specific for <i>Bp</i> LPS O polysaccharide (OPS). Unlike the typical Type A banding pattern of most <i>Bp</i> strains (1106a), the variants displayed identical higher molecular weight patterns.</p

Comparison of the virulence of variants Smooth and Rough in a mouse infection model.

<p>BALB/c mice were challenged IP with five ten-fold dilutions of Smooth (panel A) or Rough (panel B), as described previously [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0171363#pone.0171363.ref011" target="_blank">11</a>]. The decline in numbers of mice surviving with time for each dose group (10 mice/group) is shown. Infection with Rough was associated with more lethality/morbidity and a faster loss of survival than infection with Smooth.</p

Cytotoxicity of the Smooth and Rough variants of <i>Bp</i> MSHR5848 in J774.A1 macrophage cultures.

<p>Cytotoxicity of the Smooth and Rough variants of <i>Bp</i> MSHR5848 in J774.A1 macrophage cultures.</p