Microorganisms of the Upper Atmosphere: I. Instrumentation for Isokinetic Air Sampling at Altitude

An aircraft-mounted air-sampling system for the collection of microorganisms from the upper air is described. The system is highly suitable for use in upper air sampling from an aircraft because (i) it is an isokinetic system, (ii) it is simple to operate, (iii) it employs a dry filter, (iv) the volume sampled can be easily determined, and (v) it operates efficiently at all altitudes and ambient temperatures.     

Silica Gel Medium for Enumeration of Petroleumlytic Microorganisms in the Marine Environment

A silica gel medium was developed for the enumeration of petroleumlytic microorganisms in the marine environment. The medium was satisfactorily used for the investigation of the vertical distribution of bacteria in seawater from the surface to 1,000 m depth of western north Pacific central water as well as the neritic region of Japan.     

Rapid Technique for Enumeration and Isolation of Peroxidase-producing Microorganisms

Peroxidase-positive microorganisms were enumerated on agar plates by use of a p-anisidine-H(2)O(2) spray. Combined with replica-plating, the same technique allowed the selective isolation of peroxidase-producing microbial cultures.     

Agar Plate Method for Detection and Enumeration of Alkylbenzenesulfonate-Degrading Microorganisms

A simple method for detection and enumeration of alkylbenzenesulfonate (ABS)-degrading microorganisms by using agar plates was developed and used in microbiological studies of coastal marine and polluted river waters. The method depends upon the color responses of neutral red in alkaline medium. Neutral red changes from pink, when it enters into ABS micelles, to yellow, when the ABS is degraded, and does not form micelles. When neutral red-tris(hydroxymethyl)-aminomethane buffer solution and then cationic surfactant solution were sprayed onto the agar surface of ABS-nutrient agar cultures, transparent haloes appeared around the colonies of ABS-degrading microorganisms against a pink background. Viable counts of ABS-degrading bacteria isolated from both seawater and freshwater environments were considerably higher in polluted waters than in less polluted areas. Viable counts of ABS-degrading bacteria averaged 1.5 x 105/ml in samples from the surface water of polluted Tokyo Bay and 3.0 x 104/ml in samples from the surface water of polluted Tamagawa River but were fewer in number in samples from less polluted waters.     

Development of the Laryngeal Air Sac in Chimpanzees

Though many nonhuman primates possess a laryngeal sac, the great apes are unique in their great size. Though an enlarged sac probably arose in their common ancestor, its functional adaptations remain a matter of debate. Its development in extant great apes is likely to provide valuable information to clarify the issue. We used magnetic resonance imaging to examine the development of the laryngeal sac in 3 living chimpanzees, age 4 mo–5 yr, and identified 2 distinct growth phases of the sac. A gradual growth of the sac in early infancy results in a configuration so that it occupies the ventral region of the neck; many adult nonhominoid primates having a sac show the configuration. The subsequent rapid expansion of the sac in late infancy causes the final configuration in chimpanzees, wherein the sac expands into the pectoral, clavicular, and axillary regions. The latter phase possibly arose at latest in the last common ancestor of extant great apes and contributed to the evolution of the enlarged sac, despite the later evolutionary diversification in adult sac anatomy and growth. As many studies have advocated, the enlarged sac probably plays a role in vocalization in adults. However, physiological modifications in the laryngeal region during infancy are likely to provide valuable information to evaluate the functional adaptations of the enlarged sac in the great apes.     

An Immunological Assay for Detection and Enumeration of Thermophilic Biomining Microorganisms

A specific, fast, and sensitive nonradioactive immunobinding assay for the detection and enumeration of the moderate thermophile Thiobacillus caldus and the thermophilic archaeon Sulfolobus acidocaldarius was developed. It employs enhanced chemiluminescence or peroxidase-conjugated immunoglobulins in a dot or slot blotting system and is very convenient for monitoring thermophilic bioleaching microorganisms in effluents from industrial bioleaching processes.     

Sheen Screen, a Miniaturized Most-Probable-Number Method for Enumeration of Oil-Degrading Microorganisms

Sheen Screen is a miniaturized method for enumerating oil-degrading microorganisms. The technique relies on the ability of oil-degrading microorganisms to emulsify oil when provided as a sole carbon source in 24-well tissue culture plates. Sediments that actively respire hydrocarbons have high numbers of Sheen Screen-positive microorganisms.     

Simple Cannulation Procedure for Serial Blood Sampling Through Cutaneous Ulnar Vein in Chickens

The objective of the study was to collect repeated, low-stress blood samples from the ulnar vein of chickens required for pharmacokinetic studies or hormonal assays. The study used 5 apparently healthy, unsexed, commercial broiler chickens about 6 weeks old and weighing 1.7–1.9 kg for serial sampling of blood. The study prepared the birds prior to cannulation and penetrated the catheter through the skin and into the lumen of the ulnar vein. The study successfully carried out serial blood samplings in 4 of 5 cannulated birds. Heparin (10%) solution maintained patency and prevented blood clot formation inside the cannula. However, the study found repeated clotting occurring in 1 bird. Cannula failed to maintain patency; the study could not carry out blood sampling properly, which was attributed to air embolism that might have occurred during catheter manipulation or repeated filling of cannula with heparin solution. The study observed no hematoma or inflammation at the site of cannulation. Owing to the advantages and to facilitate compliance with nonhuman animal welfare, this technique seems simple and efficient, allowing adoption for serial blood collection in chickens.     

Evaluation of Methods for Sampling, Recovery, and Enumeration of Bacteria Applied to the Phylloplane

Determining the fate and survival of genetically engineered microorganisms released into the environment requires the development and application of accurate and practical methods of detection and enumeration. Several experiments were performed to examine quantitative recovery methods that are commonly used or that have potential applications. In these experiments, Erwinia herbicola and Enterobacter cloacae were applied in greenhouses to Blue Lake bush beans (Phaseolus vulgaris) and Cayuse oats (Avena sativa). Sampling indicated that the variance in bacterial counts among leaves increased over time and that this increase caused an overestimation of the mean population size by bulk leaf samples relative to single leaf samples. An increase in the number of leaves in a bulk sample, above a minimum number, did not significantly reduce the variance between samples. Experiments evaluating recovery methods demonstrated that recovery of bacteria from leaves was significantly better with stomacher blending, than with blending, sonication, or washing and that the recovery efficiency was constant over a range of sample inoculum densities. Delayed processing of leaf samples, by storage in a freezer, did not significantly lower survival and recovery of microorganisms when storage was short term and leaves were not stored in buffer. The drop plate technique for enumeration of bacteria did not significantly differ from the spread plate method. Results of these sampling, recovery, and enumeration experiments indicate a need for increased development and standardization of methods used by researchers as there are significant differences among, and also important limitations to, some of the methods used.     

Review of Some Important Techniques in Wildlife Sampling and Sampling Errors

This paper reviews some of the important methods for estimating animal numbers or densities based on (i) direct counts of population units as used in quadrat, strip, line-transect and line-intercept sampling and (ii) indirect counts and indices, such as capture-mark recapture, change-in-ratio, catch-effort methods and indices based on track counts, call, roadside and pellet-group counts, etc.     

Inexpensive Device for the Aerobic and Anaerobic Sampling of Microorganisms in Lake and Shallow Ocean Waters

An inexpensive device suitable for sampling microorganisms in water and easily constructed from readily available laboratory equipment is described. The need to transfer subsamples to culturing flasks after collection is eliminated by partly filling the sampling vessels with growth medium prior to sampling. The device is readily adapted for sampling different volumes, is simple and quick to operate, and is suitable for use with prereduced media. Contamination from layers other than that being sampled is insignificant.     

Enumeration and Diversity of Campylobacters and Bacteriophages Isolated during the Rearing Cycles of Free-Range and Organic Chickens

Campylobacters and Campylobacter-specific bacteriophages were isolated and enumerated during the rearing cycle of free-range (56 days) and organic chickens (73 days) at 3-day intervals from hatching until slaughter. In both flocks Campylobacter jejuni was the initial colonizer but Campylobacter coli was detected more frequently from 5 weeks of age. The diversity of the Campylobacter isolates was examined by pulsed-field gel electrophoresis of SmaI-digested genomic DNA and antimicrobial resistance typing. Bacteriophages were isolated from 51% (19 of 37 birds) of Campylobacter-positive organic birds (log₁₀ 2.5 to log₁₀ 5.7 PFU/g of cecal contents). The bacteriophages were all typical group III Campylobacter bacteriophages in terms of genomic size but could be characterized in terms of their host range and placed into five different groups. In contrast to the organic birds, anti-Campylobacter activity (bacteriocin-like) was observed in 26% (10 of 38 birds) of Campylobacter-positive free-range birds, and only one bacteriophage was isolated. Appearance of either bacteriophages or anti-Campylobacter activity was associated with changes in the levels of colonization and the predominant genotypes and species isolated. The frequency and potential influence of naturally occurring bacteriophages and/or inhibitory substances on the diversity and fluctuations of populations of campylobacters have not previously been reported in either free-range or organic chickens.     

Convenient Sampling of Air Bacteria in Operating Rooms

A convenient arrangement for sampling air bacteria in operating rooms with a slit sampler (the Fort Detrick sampler) is described. Its purpose is to contribute as far as possible to the convenience of the surgical staff and thereby to the safety of the patient. It has the advantages of recording minute-to-minute changes in bacterial air count; it is unobtrusive and yet can be continually observed by a technician; it is not noisy and avoids the dangers of explosion and static electricity; it is inexpensive, and parts are easily replaced; and finally it provides a means of keeping permanent photographic records of bacterial counts. Results of a preliminary trial of this method appeared to be satisfactory.     

Evaluation of Membrane Filter Field Monitors for Microbiological Air Sampling

Due to area constraints encountered in assembly and testing areas of spacecraft, the membrane filter field monitor (MF) and the National Aeronautics and Space Administration-accepted Reyniers slit air sampler were compared for recovery of airborne microbial contamination. The intramural air in a microbiological laboratory area and a clean room environment used for the assembly and testing of the Apollo spacecraft was studied. A significantly higher number of microorganisms was recovered by the Reyniers sampler. A high degree of consistency between the two sampling methods was shown by a regression analysis, with a correlation coefficient of 0.93. The MF samplers detected 79% of the concentration measured by the Reyniers slit samplers. The types of microorganisms identified from both sampling methods were similar. Variables in the MF samplers, such as pore size, relative humidity, and flow rates, have been studied, but no effect was noted on recovery. The results show that the MF method could be used to estimate the number and types of microorganisms found in the air.     

Microorganisms of the Upper Atmosphere: II. Microorganisms in Two Types of Air Masses at 690 Meters over a City

The viable micropopulation found, at altitude over a city, in a land air mass was significantly higher than that found in a marine-influenced air mass. The percentage distribution of bacteria and fungi was approximately equal in both types of air masses. This indicates that, under the conditions of the experiment, the marine air mass was influenced by the land area over which it traveled during passage from its source to the sampling area. Activities taking place within the city significantly increased the micropopulation at altitude. This increase was quantitatively so small that it was not identifiable when the micropopulation moving into the city was high-as in a land air mass-but was recognizable when the micropopulation was low-as in a marine-influenced air mass. The modification of the micropopulation at altitude by temperature inversions was shown.     

Microorganisms of the Upper Atmosphere: IV. Microorganisms of a Land Air Mass as It Traverses an Ocean

The viable micropopulation at three altitudes (152, 1,066, and 1,981 meters) of a land air mass as it traversed an ocean was determined. At the low altitude, a consistent pattern of decrease in numbers of land-originating microorganisms with increasing distance from shore was observed. At the higher altitudes, the observed pattern was one of irregularity. At the lower altitude the percentages of bacteria and fungi were approximately equal near the coast, but as distance from the coast increased, fungi predominated over the bacteria at all sampling locations. The greatest number of different genera of both bacteria and fungi were obtained at or near the coast. Fungi of the genera Alternaria, Hormodendron, Penicillium, and Aspergillus, and bacteria of the genera Micrococcus and Bacterium predominated at all altitudes and locations.     

On the Use of Jack‐Knife Techniques in Systematic Sampling

We have proposed two general classes of ratio and product type estimators to estimate an unknown population parameter of a response variable y under systematic sampling strategy. Jack‐Knife technique is employed to make the classes almost/exactly unbiased and sampling variance of the proposed estimators are derived to the first order of approximation. The merits of the proposed estimators over other estimators are discussed in this paper.     

Air Passive Sampling for the Screening of Inhalation Risks of POPs Near an Incineration Plant

The use of passive air samplers (PAS) in recent years to monitor the occurrence of persistent organic pollutants (POPs) in air has been increasing, not only globally, but also at regional and/or local scales. The present study was aimed at analyzing the airborne concentrations of the POPs: polychlorinated biphenyls (PCBs), polybrominated diphenyl ethers (PBDEs), and polychlorinated naphthalenes (PCNs) in the vicinity of a municipal solid waste incinerator (MSWI) in Tarragona (Catalonia, Spain). Mean air levels of PCBs, PBDEs, and PCNs were 44.1, 65.0, and 6.65 pg/m³, respectively. No differences were noted according to the distance or direction from the plant for any chemical. Furthermore, this information was used to estimate the air inhalation of POPs for the local population. Environmental exposure to POPs through air accounted for only <2% of the total when compared with the dietary intake. The overall results indicate that the MSWI here assessed does not mean a relevant increase on human health risks derived from inhalation of POPs. PAS can also be utilized for human health risk assessment studies, being also suitable tools to control the environmental burden of non-regulated chemicals, for which no emission thresholds have yet been derived.     

Methods for Integrated Air Sampling and DNA Analysis for Detection of Airborne Fungal Spores

Integrated air sampling and PCR-based methods for detecting airborne fungal spores, using Penicillium roqueforti as a model fungus, are described. P. roqueforti spores were collected directly into Eppendorf tubes using a miniature cyclone-type air sampler. They were then suspended in 0.1% Nonidet P-40, and counted using microscopy. Serial dilutions of the spores were made. Three methods were used to produce DNA for PCR tests: adding untreated spores to PCRs, disrupting spores (fracturing of spore walls to release the contents) using Ballotini beads, and disrupting spores followed by DNA purification. Three P. roqueforti-specific assays were tested: single-step PCR, nested PCR, and PCR followed by Southern blotting and probing. Disrupting the spores was found to be essential for achieving maximum sensitivity of the assay. Adding untreated spores to the PCR did allow the detection of P. roqueforti, but this was never achieved when fewer than 1,000 spores were added to the PCR. By disrupting the spores, with or without subsequent DNA purification, it was possible to detect DNA from a single spore. When known quantities of P. roqueforti spores were added to air samples consisting of high concentrations of unidentified fungal spores, pollen, and dust, detection sensitivity was reduced. P. roqueforti DNA could not be detected using untreated or disrupted spore suspensions added to the PCRs. However, using purified DNA, it was possible to detect 10 P. roqueforti spores in a background of 4,500 other spores. For all DNA extraction methods, nested PCR was more sensitive than single-step PCR or PCR followed by Southern blotting.     

前路及后路手术治疗高龄胸椎结核的临床疗效比较

目的:观察和比较前路及后路手术治疗高龄胸椎结核的临床疗效及其安全性.方法:选择2000年1月～2012年1月我科收治的51例高龄胸椎结核手术患者,其中18例行胸椎前路(A组),33例行胸椎后路(B组)手术治疗.观察和比较两组患者的手术时间、切口长度、出血量、COBB角度改善率、平均住院时间、并发症的发生率和术后生活质量的改善情况.结果:51例患者随访6～70个月,平均58±3个月,全部患者植骨融合,神经功能(Frankel分级)均恢复1～2级.两组患者的手术时间、切口长度、出血量无明显差异(P＞0.05).A组患者的平均住院时间为21.5± 4.9天,较B组患者(13± 3.5天)显著延长(P＜0.05);A组发生肺部感染5例,胸膜破裂并血气胸4例,泌尿系感染1例,伤口延迟愈合4例,而B组发生肺部感染例4例,伤口延迟愈合8例;A组肺部及胸腔并发症的发生率(27.8％、22.2％)较B组(12.1％、0)显著升高(P＜0.05).A组脊柱矫形效果(矫正率21.6％)显著低于B组(47.2％)(P＜0.05).A组生活质量改善20分,B组改善31分.结论:前路及后路手术治疗高龄(年龄＞70岁)老年胸椎结核均能取得良好治疗效果;与胸椎前路手术相比,胸椎后路手术可以更加有效地减少并发症,缩短住院时间,改善术后生活质量.