Home About us Contact
|
Home About us Contact | ||
|
|
||
Microscopic Counting (microscopic + counting)
Selected AbstractsThe coupling of biological iron cycling and mineral weathering during saprolite formation, Luquillo Mountains, Puerto RicoGEOBIOLOGY, Issue 4 2005H. L. BUSS ABSTRACT Corestones of quartz diorite bedrock in the Rio Icacos watershed in Puerto Rico weather spheroidally to form concentric sets of partially weathered rock layers (referred to here as rindlets) that slowly transform to saprolite. The rindlet zone (0.2,2 m thick) is overlain by saprolite (2,8 m) topped by soil (0.5,1 m). With the objective of understanding interactions between weathering, substrate availability, and resident micro-organisms, we made geochemical and microbiological measurements as a function of depth in 5 m of regolith (soil + saprolite). We employed direct microscopic counting of total cell densities; enumeration of culturable aerobic heterotrophs; extraction of microbial DNA for yield calculations; and biochemical tests for iron-oxidizing bacteria. Total cell densities, which ranged from 2.5 × 106 to 1.6 × 1010 g,1 regolith, were higher than 108 g,1 at three depths: in the upper 1 m, at 2.1 m, and between 3.7 and 4.9 m, just above the rindlet zone. High proportions of inactive or unculturable cells were indicated throughout the profile by very low percentages of culturable heterotrophs (0.0004% to 0.02% of total cell densities). The observed increases in total and culturable cells and DNA yields at lower depths were not correlated with organic carbon or total iron but were correlated with moisture and HCl-extractable iron. Biochemical tests for aerobic iron-oxidizers were also positive at 0.15,0.6 m, at 2.1,2.4 m, and at 4.9 m depths. To interpret microbial populations within the context of weathering reactions, we developed a model for estimating growth rates of lithoautotrophs and heterotrophs based on measured substrate fluxes. The calculations and observations are consistent with a model wherein electron donor flux driving bacterial growth at the saprolite,bedrock interface is dominated by Fe(II) and where autotrophic iron-oxidizing bacteria support the heterotrophic population and contribute to bedrock disaggregation and saprolite formation. [source] Hydrocarbon accumulation by picocyanobacteria from the Arabian GulfJOURNAL OF APPLIED MICROBIOLOGY, Issue 3 2001R.H. Al-Hasan Aims:,The objective of this work was to study picocyanobacteria in the Arabian Gulf water in relation to oil pollution. Methods and Results:,Epifluorescent microscopic counting showed that offshore water samples along the Kuwaiti coast of the Arabian Gulf were rich in picocyanobacteria which ranged in numbers between about 1 × 105 and 6 × 105 ml,1. Most dominant was the genus Synechococcus; less dominant genera were Synechocystis, Pleurocapsa and Dermocarpella. All isolates grew well in an inorganic medium containing up to 0·1% crude oil (w/v) and could survive in the presence of up to 1% crude oil. Hydrocarbon analysis by gas liquid chromatography (GLC) showed that representative strains of the four genera had the potential for the accumulation of hydrocarbons (the aliphatic n -hexadecane, aromatic phenanthrene and crude oil hydrocarbons) from aqueous media. Electron microscopy showed that the cells of these strains appeared to store hydrocarbons in their inter thylakoid spaces. Analysis by GLC of constituent fatty acids of total lipids and individual lipid classes from representative picoplankton strains grown in the absence and presence of hydrocarbons showed, however, that the fatty acid patterns were not markedly affected by the hydrocabon substrates, meaning that the test strains could not oxidize the accumulated hydrocarbons. Conclusions:,The Arabian Gulf is among the water bodies of the world richest in picocyanobacteria. These micro-organisms accumulate hydrocarbons from the water body, but do not biodegrade these compounds. It is assumed that hydrocarbon-utilizing bacteria that were always found associated with all picocyanobacteria in nature may carry out the biodegradation of these compounds. Significance and Importance of the Study:,The results shed light on the potential role of picocyanobacteria in controlling marine oil pollution. [source] Validation of the FACSCount AF System for Determination of Sperm Concentration in Boar SemenREPRODUCTION IN DOMESTIC ANIMALS, Issue 6 2002C Hansen Contents A flow cytometric method has been developed for rapid determination of sperm concentration in semen from various mammalian species., All cells containing DNA are stained with SYBR-14 or propidium iodide (PI) and sperm concentration is determined in relation to an internal standard of fluorescent microspheres (beads). Satisfactory staining can be achieved within 2,3 min and the following flow cytometric analysis on the FACSCount AF System rapidly provides the user with a precise and accurate assessment of the sperm concentration. In this study, the FACSCount AF System and Sperm Counting Reagent (BD Biosciences) was compared with microscopic counting using a Bürker,Türk haemocytometer. In addition, sperm concentration was determined using the Corning 254 spectrophotometer which is used routinely by Danish artificial insemination stations for boars. The results show that the agreement between flow cytometry and microscopic counting is very high. The slope for the regression line was 1.12 (SE = 0.03) with an estimated intercept with the Y-axis of 22 × 106sperm/ml (SE = 10 × 106 sperm/ml) and an estimated error of the model of 10 × 106 sperm/ml. For the spectrophotometer, the slope of the regression line was 1.09 (SE = 0.07) with an estimated intercept of 137 × 106 sperm/ml (SE = 25 × 106 sperm/ml). The average error made by the spectrophotometer was 55 × 106 sperm/ml. In addition, the results obtained using flow cytometry was highly repeatable (CV = 2.7%) in comparison with the spectrophotometric method (CV = 6.3%). These results indicate that the FACSCount AF System is a valuable tool for precise and accurate assessment of sperm concentration in boar semen and that use of this system may lead to production of more uniform insemination doses containing a specific number of sperm per dose. [source] Quantification of Chlamydia pneumoniae in cultured human macrophages and HL cells: comparison of real-time PCR, immunofluorescence and ELISA methodsAPMIS, Issue 1 2010KARI POIKONEN Poikonen K, Lajunen T, Silvennoinen-Kassinen S, Leinonen M, Saikku P. Quantification of Chlamydia pneumoniae in cultured human macrophages and HL cells: comparison of real-time PCR, immunofluorescence and ELISA methods. APMIS 2010; 118: 45,8. Chlamydia pneumoniae is an intracellular gram-negative bacterium, which replicates only in eukaryotic cells. Quantification of C. pneumoniae in cell culture is needed when studying e.g. the effect of drugs or host cell factors on infectivity and replication. Conventionally, this has been performed by immunofluorescence staining and microscopic counting of chlamydial inclusions. However, this method is usable only if the cell numbers do not fluctuate in cell culture vials and the inclusions are uniform. In macrophages, inclusions are often aberrant, their sizes vary, and multiple inclusions are also seen. Therefore, methods are needed to quantify exact amounts of C. pneumoniae in cells. Here, we describe a new method based on the real-time PCR quantification of chlamydial genomes adjusted to the number of human genomes in cultures. In human epithelial (HL) cell cultures, the C. pneumoniae inclusion numbers and the ratio of C. pneumonia genomes/human genome (Cpn/Hum) correlated significantly (r = 0.978, p < 0.001); thus with HL cells, both methods are usable. However, in macrophage cultures, the correlation was weaker (r = 0.133, p = 0.036) and we recommend PCR quantification for exact measurements. [source] | ||||||||||