Microbiology: Principles and Explorations
10th Edition
ISBN: 9781119390114
Author: Black
Publisher: WILEY
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Chapter 7, Problem 1.4SC
Why does a direct microscopic count of bacteria using a Petroff-Hausser chamber not give you a viable count? How does this differ from the spread plate and pour plate methods?
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1) both streak plating and pour plating produce isoluated colonies. What is the underlying explanation for why both methods work; that is, what are both methods doing with repect to the bacterial cells?
2) If streak plates failed to produce isolated colonies, describe two things that you could do to improve your chance of generating isolated colonies.
3) Why do we care so much about producing isolated colonies? what is an isolated colony composed of? What can you do with an isolated colony?
To determine the number of cells in a pure culture of Klebsiella pneumoniae, you have performed a serial dilution using three tubes of sterile saline (9.9 ml each). A sample of 0.1 ml from the culture was added to tube 1. Similarly, 0.1 ml from tube 1 was used to inoculate tube 2, and tube 3 was inoculated using 0.1 ml from tube 2. A nutrient agar plate was then inoculated using 0.1 ml from tube 3 and incubated overnight. The next day, 92 colonies were observed on the plate. How many cfu/ml were in the original culture? Using a Petroff-Hauser counting chamber, the number of cells in the same culture was estimated to be 8.5 • 109 cells/ml. How can you explain these results?
Given the illustration and values below, determine the concentration of the original sample. Report results in CFU/mL or colony-forming unit/milliliter. Note that in order to observe the accuracy of results, culture plates with countable colonies between 25-250 CFU are considered in standard bacterial plate count.
Chapter 7 Solutions
Microbiology: Principles and Explorations
Ch. 7 - What are the differences between the lag phase and...Ch. 7 - How does logarithmic rate of increase differ from...Ch. 7 - Prob. 1.3SCCh. 7 - Why does a direct microscopic count of bacteria...Ch. 7 - What does the ending -phile mean? Distinguish...Ch. 7 - What enzymes do most obligate anaerobes lack? How...Ch. 7 - Prob. 2.3SCCh. 7 - Prob. 3.1SCCh. 7 - Distinguish between the various kinds of media:...Ch. 7 - What is the purpose of a stock culture? Why is it...
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- There are two cultures of yeast cells in the pictures, one has been incubated for 6 hours and one has been incubated for 24 hours. After a 10x dilution by taking 100µl of each culture and adding it to 900 µl water in a microcentrifuge tube and 100µl sample from the tube was taken to view in the counting chamber. a) Count the total number of yeast cells for each culture respectively b) Calculate the concentration and density of yeast cells for each culture respectivelyarrow_forwardYou are given a bacterial culture which has a concentration of approximately 5.0 x 10^8 cells/mL. List a series of dilutions and platings that you could carry out in order to determine the exact concentration of the culture. Note that you must plate four plates from a minimum of two dilution tubes. The volumes plated should be in the range of 0.1 mL – 1.0 mL. Duplicate volumes may not be plated from any one dilution tube. Each plating should aim for a count between 30 and 300 CFUs. You can select any value from 30-300 for CFU and any volume from 0.1-1.0 to find out dilution schemearrow_forwardWhy is the plate count method a determination of the number of viable cells?arrow_forward
- A sample of water was enumerated using a counting chamber (haemocytometer) and 220 bacterial cells were counted in 25 small squares (each of volume 2.5 x 10-7 cm3). Viable counts were carried out on the same culture using both the pour plate method (incorporating 1 mL samples of a range of dilutions into plate count agar plates) and the Miles & Misra spread plate method, where 0.02 mL samples were spread on sectors of a plate count agar plate. For the pour plate count the average of triplicate samples was 178 colonies per plate of the 10-4dilution. For the Miles and Misra count the average of triplicate samples was 21.3 colonies per sector of the 10-4 dilution. Calculate the total count and the viable pour plate and spread plate counts and suggest possible reasons for the difference between the three different counts.arrow_forwardHow do we reconcile the number of colonies in the plates to the actual number of bacteria in the original sample? Count Plate Methodarrow_forwardWhy do serial dilution? Does serial dilution make it easier to count bacteria on a spread/streak plate?arrow_forward
- A serial dilution of overnight E.coli culture was performed by pipetting 1ml of a bacterial culture into a 9 ml LB medium. After this, from 10-4 and 10-5 dilution tubes 100µl were plated onto LB agar plates. Upon overnight incubation at 37°C, 200 colonies were counted in 10-4 and 22 colonies were present on 10-5 plates. How many colony-forming units were present per ml of the original culture? If the formula for CFU/ml =no. Of colonies/dilution factor*volume of culture platearrow_forwardA bacterial culture was grown for 12 hours. At 4-hour interval, the culture was sampled to determine the population of the culture, by transferring 25 ml of the suspension to 225 ml 0.90% NaCl. Three consecutive dilutions were further made by using I ml aliquot in 9 ml of 0.90% NaCl. One ml from each dilution was plated in each of duplicate plates. The following table shows the results of the plating method. 田 Sampling COUNTS 1st dilution 54; 61 2nd dilution 3rd dilution 4h dilution 1st 3; 7 0; 0 0,0 2nd TNTC TNCT 242: 233 28: 37 3rd TNTC TNTC INTC 249-246 * TNTC = Too numerous to count i. Illustrate the dilution series used and label the final dilution of each dilution. ii. Determine the bacterial count (CFU/ml) every 4 hours of incubation for 12 hours. Show all computations.arrow_forwardYou have used morphological observations and biochemical tests to identify two unknown bacterial isolates in your practical sessions, briefly describe three other techniques that can be used to identify unknown bacteria (you can use any reliable online resources). Note: These must not be based on biochemical tests (enzyme use, acid production etc) or morphological characteristics (size, shape, structures).arrow_forward
- What are the basic differences between the three plating methods? Fill up the following table. Streak Pour Spread Equipment/ Materials used in immobilizing and separating cells Purpose(s) (isolation, enumeration, both) Type of colonies (surface, subsurface, both) Advantage Disadvantagearrow_forwardIf you were using the quadrant streak plate method to plate a very dilute broth culture ( with many fewer bacteria the broth used for the plate picture here), would you expect to see single, isolated colored in quadrant 4 or quadrant3? Explain your answer.arrow_forwardA patient sample will be analyzed. You are responsible to quantify the bacteria number. The dilution series shown in the Figure ( Dilution.jpg) is prepared and 1 ml from each tube is plated. Plates were grown 24 hours resulting in colonies in each plate. Which plate you will use to quantify the bacteria in the patient sample? Why did you pick that plate? What is the dilution factor used in this series? Calculate the number of bacteria in the original patient sample. What would you do and why, if there were too many colonies to count on each plate?arrow_forward
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