저자는이 연구에 도움 식품 안전 실험실 학생들과 자원 봉사자 모두에게 감사의 말씀을 전합니다. 이 연구는 USDA 국가 통합 된 식품 안전 이니셔티브 (NIFSI) 부여 (보너스 번호 10507316)에 의해 투자되었다.

1. 표준 곡선 <ol><li> 37.5 mg의 형광 분말을 달아 6 ML 200 증거 에탄올에 용해. 200 증거 에탄올이 솔루션의 2 배 연속 희석을합니다. </li><li> 소용돌이 모두 10 초 샘플 및 370 nm에서의 흡광도를 측정한다. </li><li> 1 ㎖ 200 증거 에탄올 당 형광 분말의 양을 계산합니다. </li><li> 형광 분말의 농도에 기초하여 흡광도를 보여주기 위해 (x-축에서 370 ㎚에서의 Y 축과 흡광도 ㎎ / ml의 농도에서) 표준 곡선을 준비한다. </li></ol> 2. 형광 파우더와 델리 고기 (볼로냐) 접종 <ol><li> 깨끗한 칼을 사용하여 보드를 절단 약 100 mm 두께의 샘플로 델리 고기를 잘라. A, B 및 C라는 3 개의 시료를 준비 </li><li> 코트 샘플에 약간의 물기를 깨끗한 스폰지를 사용하여 형광 파우더로 균일하게. 균등 고기 샘플의 분말을 확산. 형광 분말 샘플 B와 C를 접종하지 마십시오. </리&gt; <li> 플라스틱 랩으로 샘플을 감싸고 &#39;A&#39;와 테이프로 B와 C 등 두 개의 샘플을 나머지로 형광 분말 코팅 샘플을 레이블. </li><li> 델리 경우 또는 실험이 실시 될 모의 부엌에서 4 ° C에서 개최 냉장고에 샘플을 놓습니다. </li></ol> 3. 모의 델리 주방 준비 <ol><li> 부엌에서 깨끗한 장갑과 70 % 에탄올 스프레이를 제공합니다. </li><li> 슬라이서 주변 13 W 전구 블랙 컴팩트 형광등을 장착합니다. </li><li> 보라고의 템플릿 역할을하는 알루미늄 호일의 5cm X 5cm 컷 아웃을 준비합니다. </li><li> 6 ㎖의 95 % 에탄올을 포함하는 15 ML 튜브를 준비합니다. </li></ol> 4. 비디오 설정 동시에 모의 델리의 모든 영역을 관찰하는 전략적 위치에있는 산 3 개의 비디오 카메라. (프로토콜 5에 설명 된대로) 슬라이서 절차를 시뮬레이션하기 전에 카메라의 전원을 켜고으로 (동시에 결과를 분석) 프로토콜 7 설명했다. 5. 교차 오염을 추적하기 위해 델리 저미는 절차 시뮬레이션 샘플 크기를 개선하고 변화를 반영, 각 기관의 임상 시험 심사위원회 프로토콜의 승인 후 모집 참가자들이 본 연구를 수행하기 위해. 참가자 또는 경험 소매 델리 환경에서 작업하지 않을 수 있습니다 대학생이 될 수 있습니다. 그들은 델리를 입력하면 각 참가자는 장갑을 착용하고 있는지 확인합니다. 가지고 검은 형광등 아래 사진은 다음에 &quot;전에&quot;연구가 완료되면 그림 &quot;후&quot;. 다음과 같이 각 참가자에게 명확한 서면 및 구두 지시 사항을 제공합니다 : <ol><li> 냉장고로 이동합니다. </li><li> &quot;A&quot;라는 레이블이 붙은 고기를 제거합니다. </li><li> 고기의 포장을 플라스틱 랩을 저장합니다. </li><li> 슬라이서의 운송 트레이에 고기를 넣습니다. </li><li> 고기 그립 고기를 고정합니다. </li><li> 전원 스위치를 켜십시오. </li><li> SLI를 조정CER 인덱스 노브 &quot;2&quot;. </li><li> 조각과는 델리 종이에 고기의 5 개 조각을 분배. </li><li> OFF 전원 스위치를 켜고 고기 압력을 해제. </li><li> &quot;A&quot;라는 레이블이 붙은 비닐 봉투에 고기 조각을 놓습니다. </li><li> 고기 &quot;A&quot;를 재 포장하고 냉장고로 돌아갑니다. </li><li> REPEAT은 &quot;B&quot;와 &quot;C&quot;라는 고기와 2-11 단계를 반복합니다. </li></ol> 6. 의 양을 형광 분말 델리 부엌에서 모의​​ 슬라이싱 (5.4 절)의 완료 후 형광 분말을 계량. 이것은 다음과 같이 경우 : <ol><li> 멸균 5cm X 5cm 알루미늄 호일 템플릿을 사용하여 그림 1에 표시된 영역에 놓습니다. </li><li> 지역을 채취, 95 % 에탄올에 담가 살균 알긴산 칼슘 면봉을 사용하여 멸균 폴리 프로필렌 튜브에 6 ㎖의 95 % 에탄올에 배치합니다. </li><li> 소용돌이 관 철저 및 370 nm에서의 흡광도를 읽을 수있는 유리 베트로 전송. </li><li> standar을 참조하십시오다음 식 사용 형광체 분말의 양 (y)를 결정하기 위해 D 곡선 : x는 흡습성이며 Y = MX + B, m은 표준 곡선의 기울기, b는 절편이다. </li><li> 데이터의 정규화를 들어, 다음 수식을 사용하여 샘​​플링 된 면적당 형광체 분말의 비율을 계산한다 : </li></ol><img src="/files/ftp_upload/51402/51402eq1.jpg" alt="포뮬러 1" /> 7. 비디오 분석 비디오 테이프 참가자들이 작업을 완료 5.1-5.12로. 동기화 및 각도가 동시에 볼 수 있도록 DVD 디스크에 비디오 레코딩을 저장한다. 손으로 건 드리면 수 델리 환경 (4 다른 연구자가 독립적으로 설정 완전한 데이터를 분석)의 다양한 표면에 (자원 봉사자 모의 델리에있는 영역을 터치하는 횟수)를 기록하는 비디오를 시청.

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관심 없음 충돌 선언하지 않습니다.

이 연구의 전반적인 목표는 세균의 교차 오염이 식중독의 위험을 관리하고 줄일 수있는 전략을 개발하기 위해 소매 델리 환경에서 발생할 수있는 방법을 설명했다. 소매 델리 환경 L.에 대한 특히 높은 위험을 가지고 오염 모노 사이토 겐. USDA-FSIS는 소매점에서 슬라이스 델리 고기 델리 고기 슬라이스 및 연방 검사 공장 9시 포장 리스테리아와 일곱 배 높은 연관성을 가지고 ?...

특히 소매 부분의 음식 준비 환경에서 인성 병원균의 교차 오염으로 인해 고기뿐만 아니라 야채 1-5 다양한 소스에서 식중독의 위험이 증가의 주요 관심사입니다. 대부분의 경우, 세균성 병원체에 오염 된 식품 (6)를 통해 소매 유통 환경을 입력합니다. 이전의 소비 7에 더 개입이나 치료 (즉, 조리 또는 가열)이 보통이 없기 때문에 소매 수준에서 준비를 먹을 준비가 된 음식은 특별한 관심의이다. 또한, 오염 RTE 식품에 존재하는 병원체는 소매 환경에서 다른 식품 또는 식품 접촉 표면에 전사 할 수있다. 교차 오염은 소매 음식 환경의 거의 모든 유형에서 발생할 수 있지만, 샌드위치 가게는 특히 때문에 병원균 리스테리아 monocytog와 RTE 델리 고기의 협회의 특정 관심의ENES 2. 미국 식품의 약국 (FDA) 및 농업 식품 안전 및 검사 서비스 (USDA-FSIS)의 미국학과에서 실시한 위험 평가 분석을 바탕으로, 오염 된 델리 고기는 리스테리아 증의 경우 90 %에 대한 가능성이 책임이 있습니다 - 희귀 L.에 의한 아직 심각한 인간의 질병 리스테리아 모노 사이토 겐 - 미국 8. 일반적으로, 소매 및 식품 서비스 시설에 슬라이스 델리 고기는 L.의 위험성 증가와 관련이있다 모노 사이토는 제조 (9)에 의해 USDA 검사 공장에서 presliced ​​가공 육류에 비해. 이 때문에 식품 취급자, 들어오는 원료 성분, 또는 후 처리 7 오염 될 수있다 가공 식품에 의한 병원균의 도입의 증가 가능성이 될 수 있습니다. 전반적으로, 리스테리아는 미국에서 매년 식중독으로 인한 예상 사망자의 약 28 %를 차지 때문에에의L.의 주름 위험 소매 수준에서 슬라이스 델리 고기에 오염 모노 사이토 겐, 델리 슬라이서 자체는 소매 환경 2,10의 인성 병원균의 표면 전송을위한 중요한 요소로 확인되었습니다. 쉰 1에 나타난 바와 같이, 슬라이스 전에 소비의 마지막 처리 단계이며, 따라서, 델리 슬라이서 이해와 교차 오염을 방지 한 모두를위한 중요 관리 점 고려되어야한다. 본 연구의 전반적인 목표는 글로 세균 같은 형광 분말을 사용 델리 소매 환경에서 오염을 추적하는 방법을 개발하는 것이었다. 이는 세균 교차 오염 및 위생 관행 11,12을 시뮬레이션하는 데 사용 된 형광 화합물이다. 이 연구는 슬라이서, 주변 환경, 기타 가공 육류에 오염 된 고기에서 병원균의 이동을 추적 할 수 있도록 설계되었습니다. 그것은 CONT의 경로를 이해하는 것이 중요합니다효과적인 개입 및 소매 직원과 관리자를위한 교육 자료를 모두 개발하는 아 민화. 유사한 접근 방식은 다른 처리 환경 (예를 들어, 가금류, 쇠고기), 농장, 병원에서 병원균의 이동을 시뮬레이션하기 위해 사용될 수있다.

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		<col />
		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr height="43">
			<td height="43" style="height:43px;width:216px;">Glo-Germ Powder</td>
			<td style="width:71px;">Glo Germ Co</td>
			<td style="width:119px;"></td>
			<td style="width:167px;">Purchased from vendor</td>
			<td style="width:64px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:216px;">Ethanol</td>
			<td style="width:71px;">Sigma</td>
			<td style="width:119px;">E7023</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:216px;">Permanent markets</td>
			<td style="width:71px;">Sharpie</td>
			<td style="width:119px;"></td>
			<td colspan="2">Purchased from stationary store</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:216px;">Gloves</td>
			<td style="width:71px;">VWR</td>
			<td style="width:119px;">82026-424</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:216px;">Deli Meat</td>
			<td style="width:71px;">NA</td>
			<td style="width:119px;">NA</td>
			<td colspan="2">Bologna Chub from regular grocery store</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:216px;">Cutting Board</td>
			<td style="width:71px;">NA</td>
			<td style="width:119px;">NA</td>
			<td colspan="2">A regukar kitchen cutting board</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:216px;">Knife</td>
			<td style="width:71px;">NA</td>
			<td style="width:119px;">NA</td>
			<td>A regular kitchen knife&#160;</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:216px;">5 x 5 cm sterile templates</td>
			<td style="width:71px;">NA</td>
			<td style="width:119px;">NA</td>
			<td colspan="2">Aluminium foil templates cut into 5x 5 cm templates and sterilized&#160;</td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:216px;">15 ml polypropylene centrifuge tubes</td>
			<td style="width:71px;">VWR</td>
			<td style="width:119px;">89039-664</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:216px;">Cotton swabs</td>
			<td style="width:71px;">Puritan</td>
			<td style="width:119px;">25-806</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:216px;">Glass cuvettes</td>
			<td style="width:71px;">VWR</td>
			<td style="width:119px;">470019-186</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:216px;">Vortex</td>
			<td style="width:71px;">VWR</td>
			<td style="width:119px;">58816-121</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:216px;">Flip camera</td>
			<td style="width:71px;">Flip Ultra HD</td>
			<td style="width:119px;">NA</td>
			<td>Purchased online</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:216px;">Deli slicer</td>
			<td style="width:71px;">Bizerba</td>
			<td style="width:119px;">SE-12</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:216px;">Deli refrigerator</td>
			<td style="width:71px;">True company</td>
			<td style="width:119px;">TDBD-722</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:216px;">Scale</td>
			<td style="width:71px;"></td>
			<td style="width:119px;">NA</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="85">
			<td height="85" style="height:85px;width:216px;">Spectrophotometer</td>
			<td style="width:71px;">Milton Roy Spectronic 20D</td>
			<td style="width:119px;">NA</td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

tracking microbial contamination in retail environments using fluorescent powder - a retail delicatessen environment example

소매 환경에서의 인성 병원균의 교차 오염이 식중독에 대한 위험이 증가에 기여하는 중요한 공중 보건 문제입니다. 이러한 델리 고기, 치즈, 경우에 신선한 농산물로 먹을 준비가 된 (RTE) 가공 식품 등 리스테리아 등 병원균의 오염으로 인한 식품 매개 질병 발생에 관여하고있다. L.에 대하여 모노 사이토는, 델리 슬라이서는 종종 교차 오염의 주요 원천입니다. 이 연구의 목적은 세균 오염을 시뮬레이션과 소매 환경이 오염을 추적하기 위해 형광 화합물을 사용하는 것이 었습니다. 모의 델리 부엌은 소매 환경을 시뮬레이션하기 위해 설계되었습니다. 델리 고기가 형광 화합물로 접종과 자원 봉사자는 식품 소매 직원의 예상과 유사한 일련의 작업을 완료하기 위해 모집했다. 자원 봉사자는 패키지를 슬라이스와 델리 냉장고에 고기를 저장하는 지시했다. 잠재적 인교차 오염은 특정 지역을 보라고하고 분광 광도계 채취 영역의 광학 밀도를 측정하여 모의 소매 환경에서 추적했다. 결과는 슬라이서의 냉장고 (즉, 델리 경우) 그립과 다양한 영역이 교차 ​​오염에 대한 높은 위험이 있다고 지적했다. 본 연구의 결과는 소매 직원 집중된 교육 자료를 개발하기 위하여 사용될 수있다. 또한, 유사한 방법 또한 식량 생산 환경 (예를 들어, 작은 농장), 병원, 요양원, 유람선 및 호텔에서 미생물 오염을 추적하는 데 사용할 수 있습니다.

그림 1은 자원 봉사자가 할당 된 작업을 완료 한 후 채취 된 슬라이서의 영역을 나타냅니다. 자원 봉사자는이 다양한 표면에 손을 접촉의 평균 주파수를 분석하기 위해 비디오 촬영을했다. 손 접촉의 빈도는 네 개의 서로 다른 관찰자에 의해 분석 및 평균 하였다. 이러한 결과는 그림 2에서 설명된다. 데이터는 고기 그립, 델리 고기 래퍼, 델리 고기, 그리고 델리 종이가 손...

Watch this Scientific Journal Video about Tracking Microbial Contamination in Retail Environments Using Fluorescent Powder - A Retail Delicatessen Environment Example at JoVE.com

Tracking Microbial Contamination in Retail Environments Using Fluorescent Powder - A Retail Delicatessen Environment Example

이 연구의 전반적인 목표는 소매 델리 설정에서 인성 병원균 리스테리아의 잠재적 교차 오염의 메커니즘을 설명했다. 이 방법론은 병원균 오염을 추적하기 위해 다른 다양한 환​​경에 적용 할 수있다.

형광 파우더를 사용하여 소매 환경에서의 미생물 오염을 추적 - 소매 델리 카트슨 환경 예

Cross contamination of foodborne pathogens in the retail environment is a significant public health issue contributing to an increased risk for foodborne ...

tracking-microbial-contamination-retail-environments-using

Research

JoVE Journal

Environment

11.0K 조회수.  University of Houston.  이 연구의 전반적인 목표는 소매 델리 설정에서 인성 병원균 리스테리아의 잠재적 교차 오염의 메커니즘을 설명했다. 이 방법론은 병원균 오염을 추적하기 위해 다른 다양한 환​​경에 적용 할 수있다. 

비디오: Tracking Microbial Contamination in Retail Environments Using Fluorescent Powder - A Retail Delicatessen Environment Example

Determination of Microbial Extracellular Enzyme Activity in Waters, Soils, and Sediments using High Throughput Microplate Assays

Much of the nutrient cycling and carbon processing in natural environments occurs through the activity of extracellular enzymes released by microorganisms. Thus, measurement of the activity of these extracellular enzymes can give insights into the rates of ecosystem level processes, such as organic matter decomposition or nitrogen and phosphorus mineralization. Assays of extracellular enzyme activity in environmental samples typically involve exposing the samples to artificial colorimetric or fluorometric substrates and tracking the rate of substrate hydrolysis. Here we describe microplate based methods for these procedures that allow the analysis of large numbers of samples within a short time frame. Samples are allowed to react with artificial substrates within 96-well microplates or deep well microplate blocks, and enzyme activity is subsequently determined by absorption or fluorescence of the resulting end product using a typical microplate reader or fluorometer. Such high throughput procedures not only facilitate comparisons between spatially separate sites or ecosystems, but also substantially reduce the cost of such assays by reducing overall reagent volumes needed per sample.

Microplate based procedures are described for the colorimetric or fluorometric analysis of extracellular enzyme activity. These procedures allow for the rapid assay of such activity in large numbers of environmental samples within a manageable time frame.

높은 처리량 마이크로 분석 실험을 사용하여 워터스, 토양 및 퇴적물에있는 미생물 세포 외 효소 활성의 결정

Much of the nutrient cycling and carbon  processing in natural environments occurs through the activity of extracellular  enzymes released by ...

연구

환경과학

Technique for Studying Arthropod and Microbial Communities within Tree Tissues

Phloem tissues of pine are habitats for many thousands of organisms. Arthropods and microbes use phloem and cambium tissues to seek mates, lay eggs, rear young, feed, or hide from natural enemies or harsh environmental conditions outside of the tree. Organisms that persist within the phloem habitat are difficult to observe given their location under bark. We provide a technique to preserve intact phloem and prepare it for experimentation with invertebrates and microorganisms. The apparatus is called a &#8216;phloem sandwich&#8217; and allows for the introduction and observation of arthropods, microbes, and other organisms. This technique has resulted in a better understanding of the feeding behaviors, life-history traits, reproduction, development, and interactions of organisms within tree phloem. The strengths of this technique include the use of inexpensive materials, variability in sandwich size, flexibility to re-open the sandwich or introduce multiple organisms through drilled holes, and the preservation and maintenance of phloem integrity. The phloem sandwich is an excellent educational tool for scientific discovery in both K-12 science courses and university research laboratories.

We provide a technique to preserve intact tree phloem and prepare it for observation. We create an apparatus called a phloem sandwich that allows for the introduction and observation of arthropods, microbes, and other organisms that inhabit phloem tissues.

나무 조직 내에서 절지 동물과 미생물 군집 유학을위한 기술

Phloem tissues of pine are habitats for many thousands of organisms.  Arthropods and microbes use phloem and cambium tissues to seek mates, lay eggs, rear ...

An Aquatic Microbial Metaproteomics Workflow: From Cells to Tryptic Peptides Suitable for Tandem Mass Spectrometry-based Analysis

Meta-omic technologies such as metagenomics, metatranscriptomics and metaproteomics can aid in the understanding of microbial community structure and metabolism. Although powerful, metagenomics alone can only elucidate functional potential. On the other hand, metaproteomics enables the description of the expressed in situ metabolism and function of a community. Here we describe a protocol for cell lysis, protein and DNA isolation, as well as peptide digestion and extraction from marine microbial cells collected on a cartridge filter unit (such as the Sterivex filter unit) and preserved in an RNA stabilization solution (like RNAlater). In mass spectrometry-based proteomics studies, the identification of peptides and proteins is performed by comparing peptide tandem mass spectra to a database of translated nucleotide sequences. Including the metagenome of a sample in the search database increases the number of peptides and proteins that can be identified from the mass spectra. Hence, in this protocol DNA is isolated from the same filter, which can be used subsequently for metagenomic analysis.

This protocol is for the extraction and concentration of protein and DNA from microbial biomass collected from seawater, followed by the generation of tryptic peptides suitable for tandem mass spectrometry-based proteomic analysis.

수생 미생물 Metaproteomics 워크 플로우 : 탠덤 질량 분석 기반 분석에 적합 트립신 펩티드로 세포

Meta-omic technologies such as metagenomics, metatranscriptomics and metaproteomics can aid in the understanding of microbial community structure and ...

Extraction and Analysis of Microbial Phospholipid Fatty Acids in Soils

Phospholipid fatty acids (PLFAs) are key components of microbial cell membranes. The analysis of PLFAs extracted from soils can provide information about the overall structure of terrestrial microbial communities. PLFA profiling has been extensively used in a range of ecosystems as a biological index of overall soil quality, and as a quantitative indicator of soil response to land management and other environmental stressors.
The standard method presented here outlines four key steps: 1. lipid extraction from soil samples with a single-phase chloroform mixture, 2. fractionation using solid phase extraction columns to isolate phospholipids from other extracted lipids, 3. methanolysis of phospholipids to produce fatty acid methyl esters (FAMEs), and 4. FAME analysis by capillary gas chromatography using a flame ionization detector (GC-FID). Two standards are used, including 1,2-dinonadecanoyl-sn-glycero-3-phosphocholine (PC(19:0/19:0)) to assess the overall recovery of the extraction method, and methyl decanoate (MeC10:0) as an internal standard (ISTD) for the GC analysis.

Phospholipid fatty acids provide information about the structure of soil microbial communities. We present methods for extraction from soil samples with a single-phase chloroform mixture, fractionation of extracted lipids using solid phase extraction columns, and methanolysis to produce fatty acid methyl esters, which are analyzed by capillary gas chromatography.

추출 및 토양의 미생물 인지질 지방산의 분석

Phospholipid fatty acids (PLFAs) are key components of microbial cell membranes. The analysis of PLFAs extracted from soils can provide information about ...

Sampling, Sorting, and Characterizing Microplastics in Aquatic Environments with High Suspended Sediment Loads and Large Floating Debris

The ubiquitous presence of plastic debris in the ocean is widely recognized by the public, scientific communities, and government agencies. However, only recently have microplastics in freshwater systems, such as rivers and lakes, been quantified. Microplastic sampling at the surface usually consists of deploying drift nets behind either a stationary or moving boat, which limits the sampling to environments with low levels of suspended sediments and floating or submerged debris. Previous studies that employed drift nets to collect microplastic debris typically used nets with &#8805;300 &#181;m mesh size, allowing plastic debris (particles and fibers) below this size to pass through the net and elude quantification. The protocol detailed here enables: 1) sample collection in environments with high suspended loads and floating or submerged debris and 2) the capture and quantification of microplastic particles and fibers &lt;300 &#181;m. Water samples were collected using a peristaltic pump in low-density polyethylene (PE) containers to be stored before filtering and analysis in the lab. Filtration was done with a custom-made microplastic filtration device containing detachable union joints that housed nylon mesh sieves and mixed cellulose ester membrane filters. Mesh sieves and membrane filters were examined with a stereomicroscope to quantify and separate microplastic particulates and fibers. These materials were then examined using a micro-attenuated total reflectance Fourier transform infrared spectrometer (micro ATR-FTIR) to determine microplastic polymer type. Recovery was measured by spiking samples using blue PE particulates and green nylon fibers; percent recovery was determined to be 100% for particulates and 92% for fibers. This protocol will guide similar studies on microplastics in high velocity rivers with high concentrations of sediment. With simple modifications to the peristaltic pump and filtration device, users can collect and analyze various sample volumes and particulate sizes.

Most microplastic research to date has occurred in marine systems where suspended solid levels are relatively low. Focus is now shifting to freshwater systems, which may feature high sediment loads and floating debris. This protocol addresses collecting and analyzing microplastic samples from aquatic environments that contain high suspended solid loads.

샘플링, 정렬, 및 높은 정지 퇴적 부하와 큰 부동 파편 수생 환경에서 Microplastics

The ubiquitous presence of plastic debris in the ocean is widely recognized by the public, scientific communities, and government agencies. However, only ...

Detection of 3-Nitrotyrosine in Atmospheric Environments via a High-performance Liquid Chromatography-electrochemical Detector System

3-nitrotyrosine (3-NT) is generated from the tyrosine residue in atmospheric bio-aerosol proteins via a reaction with ozone (O3) and nitrogen dioxide (NO2). Stable 3-NT is a specific marker for oxidative damage and is reported to have a promotive effect to elicit allergies. In the present study, we report the development of a highly sensitive assay to quantify 3-NT in air sampler filters to collect &#60; 2.5 &#181;m of particulate matter (PM2.5) from urban environmental air, including bio-aerosol. In this method, a 6 mm-diameter round hole was cut from the filters of air samplers and mixed with a nonspecific protease cocktail in order to hydrolyze proteins. Protein samples digested to the amino acid level were tested for 3-NT using a high-performance liquid chromatography-electrochemical detector (HPLC-ECD). The maximum 3-NT content was detected in a prefilter for PM of sizes from 4.5 to 7.3 &#956;m, with a detection limit of 1.13 pg/m3.

Detection of 3-Nitrotyrosine in Atmospheric Environments via a High-performance Liquid Chromatography-electrochemical Detector System

We present a method to detect 3-nitrotyrosine chemical modifications of atmospheric proteins with 6 mm-diameter rounds cut from air sampler prefilters using a high-performance liquid chromatography-electrochemical detector (HPLC-ECD).

3-Nitrotyrosine 대기 환경을 통해 고성능 액체 착 색 인쇄기-전기 화학 검출기 시스템에서의 탐지

3-nitrotyrosine (3-NT) is generated from the tyrosine residue in atmospheric bio-aerosol proteins via a reaction with ozone (O3) and nitrogen dioxide ...

Fluorescently Labeled Bacteria as a Tracer to Reveal Novel Pathways of Organic Carbon Flow in Aquatic Ecosystems

Elucidating trophic interactions, such as predation and its effects, is a frequent task for many researchers in ecology. The study of microbial communities has many limitations, and determining a predator, prey, and predatory rates is often difficult. Presented here is an optimized method based on the addition of fluorescently labelled prey as a tracer, which allows for reliable quantitation of the grazing rates in aquatic predatory eukaryotes and estimation of nutrient transfer to higher trophic levels.

Presented here is a protocol for a single-cell, epifluorescence microscopy-based technique to quantify grazing rates in aquatic predatory eukaryotes with high precision and taxonomic resolution.

수생 생태계에서 유기 탄소 흐름의 새로운 경로를 밝히기 위해 추적자로 형광으로 분류 된 박테리아

Elucidating trophic interactions, such as predation and its effects, is a frequent task for many researchers in ecology. The study of microbial ...

Enrichment and Detection of Clostridium perfringens Toxinotypes in Retail Food Samples

Clostridium perfringens (C. perfringens) is a prolific toxin producer and causes a wide range of diseases in various hosts. C. perfringens is categorized into five different toxinotypes, A through E, based on the carriage of four major toxin genes. The prevalence and distribution of these various toxinotypes is understudied, especially their pervasiveness in American retail food. Of particular interest to us are the type B and D strains, which produce epsilon toxin, an extremely lethal toxin suggested to be the environmental trigger of multiple sclerosis in humans. To evaluate the presence of different C. perfringens toxinotypes in various food samples, we developed an easy method to selectively culture these bacteria without the use of an anaerobic container system only involving three culturing steps. Food is purchased from local grocery stores and transported to the laboratory under ambient conditions. Samples are minced and inoculated into modified rapid perfringens media (RPM) and incubated overnight at 37 &#176;C in a sealed, airtight conical tube. Overnight cultures are inoculated onto a bottom layer of solid Tryptose Sulfite Cycloserine (TSC) agar, and then overlaid with a top layer of molten TSC agar, creating a &#34;sandwiched&#34;, anaerobic environment. Agar plates are incubated overnight at 37 &#176;C and then evaluated for appearance of black, sulfite-reducing colonies. C. perfringens-suspected colonies are removed from the TSC agar using sterile eye droppers, and inoculated into RPM and sub-cultured overnight at 37 &#176;C in an airtight conical tube. DNA is extracted from the RPM subculture, and then analyzed for the presence of C. perfringens toxin genes via polymerase chain reaction (PCR). Depending on the type of food sampled, typically 15&#8211;20% of samples test positive for C. perfringens.

Enrichment and Detection of Clostridium perfringens Toxinotypes in Retail Food Samples

The objective of this protocol is to detect different Clostridium perfringens toxinotypes in locally purchased foods, particularly epsilon toxin producing strain types B and D, without the use of anaerobic chambers.

소매 식품 시료에서 클로스트리디움 퍼프린지 독시변의 농축 및 검출

Clostridium perfringens (C. perfringens) is a prolific toxin producer and causes a wide range of diseases in various hosts. C. perfringens is categorized ...

In Situ Chemotaxis Assay to Examine Microbial Behavior in Aquatic Ecosystems

Microbial behaviors, such as motility and chemotaxis (the ability of a cell to alter its movement in response to a chemical gradient), are widespread across the bacterial and archaeal domains. Chemotaxis can result in substantial resource acquisition advantages in heterogeneous environments. It also plays a crucial role in symbiotic interactions, disease, and global processes, such as biogeochemical cycling. However, current techniques restrict chemotaxis research to the laboratory and are not easily applicable in the field. Presented here is a step-by-step protocol for the deployment of the in situ chemotaxis assay (ISCA), a device that enables robust interrogation of microbial chemotaxis directly in the natural environment. The ISCA is a microfluidic device consisting of a 20 well array, in which chemicals of interest can be loaded. Once deployed in aqueous environments, chemicals diffuse out of the wells, creating concentration gradients that microbes sense and respond to by swimming into the wells via chemotaxis. The well contents can then be sampled and used to (1) quantify strength of the chemotactic responses to specific compounds through flow cytometry, (2) isolate and culture responsive microorganisms, and (3) characterize the identity and genomic potential of the responding populations through molecular techniques. The ISCA is a flexible platform that can be deployed in any system with an aqueous phase, including marine, freshwater, and soil environments.

Presented here is the protocol for an in situ chemotaxis assay, a recently developed microfluidic device that enables studies of microbial behavior directly in the environment.

수생 생태계에서 미생물 행동을 검사하는 시투 화학 분석분석

Microbial behaviors, such as motility and chemotaxis (the ability of a cell to alter its movement in response to a chemical gradient), are widespread ...

Evaluating the Impact of Hydraulic Fracturing on Streams using Microbial Molecular Signatures

Hydraulic fracturing (HF), commonly called &#34;fracking&#34;, uses a mixture of high-pressure water, sand, and chemicals to fracture rocks, releasing oil and gas. This process revolutionized the U.S. energy industry, as it gives access to resources that were previously unobtainable and now produces two-thirds of the total natural gas in the United States. Although fracking has had a positive impact on the U.S. economy, several studies have highlighted its detrimental environmental effects. Of particular concern is the effect of fracking on headwater streams, which are especially important due to their disproportionately large impact on the health of the entire watershed. The bacteria within those streams can be used as indicators of stream health, as the bacteria present and their abundance in a disturbed stream would be expected to differ from those in an otherwise comparable but undisturbed stream. Therefore, this protocol aims to use the bacterial community to determine if streams have been impacted by fracking. To this end, sediment, and water samples, from streams near fracking (potentially impacted) and upstream or in a different watershed of fracking activity (unimpacted) must be collected. Those samples are then subjected to nucleic acid extraction, library preparation, and sequencing to investigate microbial community composition. Correlational analysis and machine learning models can subsequently be employed to identify which features are explanative of variation in the community, as well as identification of predictive biomarkers for fracking&#39;s impact. These methods can reveal a variety of differences in the microbial communities among headwater streams, based on the proximity to fracking, and serve as a foundation for future investigations on the environmental impact of fracking activities.

Here, we present a protocol to investigate the impacts of hydraulic fracturing on nearby streams by analyzing their water and sediment microbial communities.

미생물 분자 서명을 사용하여 스트림에 유압 골절의 영향 평가

Hydraulic fracturing (HF), commonly called &#34;fracking&#34;, uses a mixture of high-pressure water, sand, and chemicals to fracture rocks, releasing oil ...