scholarly journals Agar Desiccation – The Causes and How to Address Them

2021 ◽  
Author(s):  
Tammy Hassel
Keyword(s):  
Moisture Content ◽  
Environmental Monitoring ◽  
Culture Media ◽  
False Negative ◽  
Data Gathering ◽  
Petri Dish ◽  
Solid Culture ◽  
Critical Data ◽  
Negative Results ◽  
The Media

Solid culture media, or agar plates, are prone to desiccation. The combination of warm, dry air and limited moisture content to achieve a solid finish leads to inevitable drying out and desiccation. Cracks in the media, a loss of volume or shrinkage away from the sides of the Petri dish are evidence of desiccation. Overall, this reduces the capability of the media to support the growth of organisms. The reduction in growth support capability is particularly so if the microorganisms deposited on the surface of the media are stressed. Why is Desiccation an Issue It is a well-known fact that microbes have three basic requirements for growth; a food source, warmth and moisture. Reducing the moisture content risks inactivating and or inhibiting growth. The purpose of monitoring for microbes through the use of solid media is to capture microorganisms from the manufacturing environment and create favourable conditions to support their optimum growth, thus allowing them to replicate and form visible colonies for inspection. Viable environmental monitoring is a challenging process to optimise for; you are looking for the chance of capturing one of the very few microorganisms present in your extensive manufacturing facility. To do so, you use solid culture media to take samples, snapshots in time, of your environment. Firstly, you have a vast space to monitor, and your monitoring activity needs to find the very few microorganisms present. If you have set your monitoring programme up suitably and you successfully capture organism present in the environment but then add desiccation you risk losing critical data. Desiccation leads to a reduction in these favourable conditions and potentially a loss of critical data and false-negative results. Desiccation of media is considered a significant quality issue as it can lead to the inhibition of growth and or cell death. The action of drying out of the media can lead to the formation of a skin on the surface. The skin formation inhibits the recovery of organisms, through «bounce» or «air bounce». As the purpose of environmental monitoring is to capture and support the growth of contaminants, this issue will result in inconclusive data gathering. Desiccation is an inevitability. Cleanroom environments are by their very nature, high airflow facilities which, as you will see below, is a primary reason for the loss of moisture from agar plates. It is, therefore, important to ensure media qualification studies include a review that post use, your media, can meet the 70% growth rate recovery as specified in USP <1227>.

Fluorescent antibody techniques have allowed for the direct identification and enumeration of individual bacteria in environmental samples without requiring prior growth in culture media (Bahlool and Schmidt 1980, Cloete and Steyn 1988, Macario et al. 1989). The technique involves the use of specific antibodies raised against surface markers of defined pure cultures that are either labelled directly with fluorescent dye molecules or via a fluorescent secondary antibody. This approach has yielded important insights into the spatial distribution of microorganisms, but it suffers from a number of disadvantages. For example, expression of the antigen may be influenced by environmental factors; false-positive and false-negative results may be obtained due to cross-reactivity or lack of reaction; non-specific binding of antibodies may result in high levels of background fluorescence; and production of specific antibodies requires a pure culture of the organism of interest (Cloete and de Bruyn Various recombinant DNA techniques have subsequently been developed that are independent of cultivation methods (Fig. 1). These techniques provide ways of detecting and quantifying specific phylogenetic groups of microbes on 16S rDNA sequences, and relevant structural genes provide ways of monitoring microbial populations of environmental and industrial systems. In addition to these tools, a number of emerging technologies such as the use of biomarker genes are being increasingly used to monitor with great precision and accuracy the behaviour of microbes in the environment.

2003 ◽  
pp. 218-219
Keyword(s):  
Recombinant Dna ◽  
Culture Media ◽  
Specific Binding ◽  
Fluorescent Antibody ◽  
False Negative ◽  
Cross Reactivity ◽  
Phylogenetic Groups ◽  
Specific Antibodies ◽  
Negative Results ◽  
Recombinant Dna Techniques

Survival of Escherichia coli O157:H7 and Campylobacter jejuni in Bottled Purified Drinking Water under Different Storage Conditions

2011 ◽  
Vol 74 (2) ◽  
pp. 254-260 ◽  
Author(s):  
HAMZAH M. AL-QADIRI ◽  
XIAONAN LU ◽  
NIVIN I. AL-ALAMI ◽  
BARBARA A. RASCO
Keyword(s):  
Escherichia Coli ◽  
Drinking Water ◽  
Campylobacter Jejuni ◽  
Culture Media ◽  
False Negative ◽  
Storage Conditions ◽  
Escherichia Coli O157 ◽  
E Coli ◽  
Log Reduction ◽  
Negative Results

Survival of Escherichia coli O157:H7 and Campylobacter jejuni that were separately inoculated into bottled purified drinking water was investigated during storage at 22, 4, and −18°C for 5, 7, and 2 days, respectively. Two inoculation levels were used, 1 and 10 CFU/ml (102 and 103 CFU/100 ml). In samples inoculated with 102 CFU/100 ml, C. jejuni was not detectable (&gt;2-log reduction) after storage under the conditions specified above. E. coli O157:H7 was detected on nonselective and selective media at log reductions of 1.08 to 1.25 after storage at 22°C, 1.19 to 1.56 after storage at 4°C, and 1.54 to 1.98 after storage at −18°C. When the higher inoculation level of 103 CFU/100 ml was used, C. jejuni was able to survive at 22 and 4°C, with 2.25- and 2.17-log reductions, respectively, observed on nonselective media. At these higher inoculation levels, E. coli O157:H7 was detectable at 22, 4, and −18°C, with log reductions of 0.76, 0.97, and 1.21, respectively, achieved on nonselective media. Additionally, E. coli O157:H7 showed significant differences in culturability (P &lt; 0.05) on the nonselective and selective culture media under the different storage conditions, with storage at −18°C for 2 days being the treatment most inhibiting. The percentage of sublethal injury of E. coli O157:H7 ranged from ~33 to 75%, indicating that microbial examination of bottled water must be done carefully, otherwise false-negative results or underestimation of bacterial numbers could pose a health risk when low levels of pathogens are present.

scholarly journals Fecundity of inbred fruit fly Drosophila melanogaster on different solid culture media: An analysis

2018 ◽  
Vol 10 (4) ◽  
pp. 1109-1114 ◽  
Author(s):  
Animesh Kumar Mohapatra ◽  
Priyamvada Pandey
Keyword(s):  
Drosophila Melanogaster ◽  
First Generation ◽  
Culture Media ◽  
Fruit Fly ◽  
Control Medium ◽  
Solid Culture ◽  
Pupal Development ◽  
Multiple Comparison Test ◽  
The Media ◽  
And Control

In the present study, wild-type Drosophila melanogaster collected from stock culture were sub-cultured in three different types of solid culture media  (corn, barley and wheat) and control medium for two weeks to produce F1 generation. The duration of larval and pupal development, number of pupal cases and hatched flies were scored for first generation. The results were analyzed by using one-way ANOVA, Bonferroni multiple comparison test and paired sample t-test. The control medium showed no pupal cases and hatched flies. Among all the three solid culture media tested, corn meal, barley meal and wheat meal, the latter showed highly significant results at p?0.001 than others. However, this parameter was not affected by the carbohydrate amount in the media. The present investigation is an attempt to evaluate the influence of different formulated solid culture media on the life span and reproduction of fruit flies.

scholarly journals APLIKASI SITOKININ TIPE PURIN DAN UREA PADA MULTIPLIKASI TUNAS ANIS (Pimpinellla anisum L.) IN VITRO

2020 ◽  
Vol 13 (1) ◽  
pp. 1
Author(s):  
OTIH ROSTIANA
Keyword(s):  
Essential Oils ◽  
Culture Media ◽  
Shoot Multiplication ◽  
Tropical Region ◽  
Herbaceous Plant ◽  
Solid Culture ◽  
Pimpinella Anisum ◽  
The Media ◽  
Initiation Stage

ABSTRAK<br />Anis (Pimpinella anisum L.) merupakan tanaman herba tahunan<br />yang termasuk ke dalam famili Umbelliferae. Buahnya diketahui<br />mengandung minyak atsiri yang didominasi senyawa trans-anethol (90%)<br />dan berkhasiat sebagai antiseptik, antispasmodik, antikanker, karminatif,<br />pelega tenggorokan, obat bronkitis, serta digunakan dalam pembuatan<br />sabun, parfum, pasta gigi, juga krim kulit. Sebagai tanaman bernilai<br />ekonomi, upaya perbanyakan anis perlu dilakukan. Perbanyakan secara in<br />vitro dengan teknik kultur jaringan merupakan salah satu metode alternatif<br />yang dapat digunakan untuk menghasilkan bibit dalam jumlah banyak,<br />seragam dan dalam waktu yang relatif singkat. Dengan penambahan<br />sitokinin sintetik tipe urea seperti thidiazuron (TDZ) dan tipe purin seperti<br />benzil amino purin (BAP) akan memacu inisiasi dan proliferasi tunas.<br />Penelitian ini bertujuan mendapatkan media yang tepat untuk menginduksi<br />tunas anis yang optimal dengan penambahan BAP atau TDZ, mengetahui<br />respon pertumbuhan dan penampakan kultur akibat penambahan berbagai<br />konsentrasi BAP atau TDZ, serta mempelajari sinergisme yang terjadi<br />antara keduanya. Pada tahap inisiasi, eksplan berupa tunas pucuk diinduksi<br />di dalam media MS padat dengan penambahan BAP (0,1 mg/l; 0,2 mg/l;<br />0,3 mg/l; 1 mg/l; 2 mg/l; 3 mg/l), atau TDZ dengan kisaran konsentrasi<br />yang sama. Tunas terbanyak yang dihasilkan dari dua jenis sitokinin pada<br />tahap ini disubkultur ke dalam media yang ditambahkan jenis sitokinin<br />yang berbeda (TDZ ke BAP atau BAP ke TDZ) pada konsentrasi 0,3 mg/l<br />atau 3 mg/l. Pada media yang ditambahkan TDZ dihasilkan tunas anis<br />lebih banyak (3,62-6,28) dibandingkan pada media yang ditambahkan<br />BAP (1,86-2,78), tetapi tunas yang dihasilkan pendek (roset). Sedangkan<br />tunas yang dihasilkan dalam media yang ditambahkan BAP beruas lebih<br />tinggi tetapi jumlah tunasnya sedikit. Subkultur tunas anis ke dalam media<br />yang diperkaya dengan sitokinin yang berbeda meningkatkan jumlah tunas<br />yang berproliferasi dan memperbaiki visual tunas.<br />Kata kunci: Anis, Pimpinellla anisum L. ,  minyak atsiri, multiplikasi tunas,<br />in vitro, TDZ, BAP, Jawa Barat<br />ABSTRACT<br />Application of purine and urea types of cytokinins in<br />shoot multiplication of Anise (Pimpinella anisum L.) in<br />vitro<br />Pimpinella anisum L. or sweet anise is an annual–herbaceous plant<br />belongs to the Umbelliferae family. The fruit of anise contains of essential<br />oil, which is mainly consisted of trans-anethol (90%). Essential oils of<br />anise is mainly used as an antiseptic, antispasmodic, anticancer,<br />carminative, expectorant and has also been used as component in soap,<br />perfumery, tooth paste, and skin cream productions. Since this crop is<br />mainly cultivated in sub tropical region, anise cultivation in Indonesia has<br />not been performed. To obtain sufficient numbers of anise planting<br />materials in vitro propagation was conducted by applying benzyl amino<br />purine (BAP) and thidiazuron (TDZ). In this research TDZ or BAP were<br />applied at various concentrations (0,1 mg/l: 0.2 mg/l; 0.3 mg/l; 1 mg/l; 2<br />mg/l; 3 mg/l), to induce shoots in MS-solid culture media. The highest<br />number of shoots obtained in those two type of cytokinins containing<br />media from the initiation stage were subcultured into the media<br />supplemented with different cytokinins (TDZ to BAP or BAP to TDZ) at<br />0.3 mg/l or 3 mg/l levels. The results showed that medium with the<br />addition of TDZ resulted in higher numbers of shoot (3,26-6,28) than that<br />of medium with an addition of BAP (1,86-2,78). However, rosette shoots<br />were dominant in TDZ containing medium. On the other hand, medium<br />with an addition of BAP resulted in less numbers of shoots with taller<br />nodes. Subculture of anise into different kinds of cytokinins increased the<br />numbers of proliferated-shoots and recovered the abnormal shoots.<br />Key words : Anise, Pimpinellla anisum L, essential oils, shoots<br />multiplication, in vitro, TDZ, BAP, West Java

A Retrospective Analysis of Joint Cultures: Only Time Will Tell (Or Will It?)

2020 ◽  
Vol 154 (Supplement_1) ◽  
pp. S138-S138
Author(s):  
D Altenburger ◽  
S Strayer ◽  
S Carroll
Keyword(s):  
Retrospective Analysis ◽  
Propionibacterium Acnes ◽  
Culture Media ◽  
False Negative ◽  
Positive Culture ◽  
Growth Time ◽  
Average Growth ◽  
Negative Results ◽  
False Negative Results ◽  
Actinomyces Species

Abstract Introduction/Objective Multiple textbooks and guidelines, including CLSI, recommend holding joint cultures for 14 days. This prevents false negative results from “slower growers” which include Actinomyces species and Propionibacterium acnes. Holding of these cultures can be a burden for small microbiology labs. Methods A retrospective analysis of three years of joint cultures held for 14 days was reviewed. The length of time for the culture to turn positive and the organism that grew was noted. Results A total of 67 positive joint cultures were evaluated, which covered a period of 3 years at our institution. Of these, only 1 culture turned positive following day 5 (Propionibacterium acnes grew on day 6). The average time to positive for Propionibacterium acnes was 4 days. In comparison, the average growth time for Staphylococcus aureus and Proteus was 1 day, Pseudomonas aeruginosa was 1.2 days, all Streptococcus species was 2.4 days and Staphylococcus epidermidis was 2.7 days. No cultures grew Actinomyces during our study. Conclusion The consequences of missing a positive culture can be grave. In general, culture media has improved and provided increased growth rates. With resources becoming more limited, it may be time to revisit guidelines. This study is too small to be conclusive; a larger study may provide confirmation of the findings.

Influence of cultivation methods on the chemical and nutritional characteristics of Lentinula edodes

2019 ◽  
pp. 1006
Author(s):  
Fabiane Bach, Cristiane Vieira Helm ◽  
Edson Alves De Lima, Marcelo Barba Bellettini, Charles Windsnon Isidoro Haminiuk
Keyword(s):  
Moisture Content ◽  
Lentinula Edodes ◽  
Culture Media ◽  
Lignin Content ◽  
C:N Ratio ◽  
Edible Mushroom ◽  
Nutritional Composition ◽  
Quercus Acutissima ◽  
Plastic Bags ◽  
The Media

Abstract: Lentinula edodes (Shiitake) is an edible mushroom with excellent nutritional potential, aroma and flavor. It’s traditionally produced on wood logs, but this practice has been replaced by cultivation on axenic substrates (AS) made from different materials that are stored in plastic bags. This papper aimed to evaluate the nutritional composition of L. edodes grown on Quercus acutíssima (QA) and AS by correlating their chemical composition with the media on which they were grown. Culture media were analyzed for their density, moisture content, ash, extractives, lignin (soluble and insoluble) and holocelulose before L. edodes inoculation, as well as after the second consecutive harvest this mushrooms. The mushrooms from the second harvest of each culture media were characterized regarding their moisture content, protein, ash, lipids, dietary fiber (soluble and insoluble), α, β and glucanas total, carbohydrates and minerals. Mushrooms cultivated on AS showed higher protein content, macrominerals and lipids, when compared to mushrooms cultivated on QA. AS initially contained lower lignin content, less holocelulose and a reduced C:N ratio when compared to QA. The results showed that the composition of the culture medium influenced the nutritional composition of L. edodes mushrooms.

scholarly journals ON A METHOD OF ISOLATING AND IDENTIFYING BACILLUS TYPHOSUS, BASED ON A STUDY OF BACILLUS TYPHOSUS AND MEMBERS OF THE COLON GROUP IN SEMI-SOLID CULTURE MEDIA

1897 ◽  
Vol 2 (6) ◽  
pp. 677-700 ◽  
Author(s):  
Philip Hanson Hiss
Keyword(s):  
Typhoid Fever ◽  
Culture Media ◽  
Tap Water ◽  
Bacterial Species ◽  
Practical Application ◽  
Solid Culture ◽  
The Media ◽  
Semi Solid

Semi-solid culture media, and more especially media rendered semi-solid by temperatures of from 30° to 40° C., seem to have an important bearing in the differentiation of bacterial species, particularly those presenting various degrees of motility. In such media not only the effect of differences in consistence on the motility of an organism may be noted, but the effect produced by various chemicals and nutrient ingredients on the growth and motility may be readily observed. By systematically varying the constituents of such media it has been possible to produce a medium in which the behavior of Bacillus typhosus differentiates it from the various members of the colon group; and also to produce a medium in which the colonies of Bacillus typhosus assume a form which distinguishes them from the colonies of the colon bacilli in plate cultures. Bacillus typhosus alone of all the organisms investigated during these experiments has displayed both the power of giving rise to thread-forming colonies in the plating medium and that of the uniform clouding of the tube medium, hence these two characters may prove to be of great value in the identification of this organism. The practical application of the use of these media has led to the ready detection of Bacillus typhosus and its isolation from the stools of patients suffering from typhoid fever. No suspected water has been subjected to test, but from the investigation of artificially infected tap-water the media here described may be assumed to have an application in the detection of Bacillus typhosus in such waters.

Critical Evaluation of Impedance Phlebography for the Diagnosis of Deep Vein Thrombosis

1974 ◽  
Vol 31 (02) ◽  
pp. 273-278
Author(s):  
Kenneth K Wu ◽  
John C Hoak ◽  
Robert W Barnes ◽  
Stuart L Frankel
Keyword(s):  
Deep Vein Thrombosis ◽  
False Positive ◽  
False Negative ◽  
Critical Evaluation ◽  
Original Method ◽  
Vein Thrombosis ◽  
Negative Results ◽  
False Negative Results ◽  
Deep Vein ◽  
Positive Results

SummaryIn order to evaluate its daily variability and reliability, impedance phlebography was performed daily or on alternate days on 61 patients with deep vein thrombosis, of whom 47 also had 125I-fibrinogen uptake tests and 22 had radiographic venography. The results showed that impedance phlebography was highly variable and poorly reliable. False positive results were noted in 8 limbs (18%) and false negative results in 3 limbs (7%). Despite its being simple, rapid and noninvasive, its clinical usefulness is doubtful when performed according to the original method.

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