Food Microbiology and Safety - Science topic

Hi Twana,

The total mesophilic plant count will be unsuitable in case of fermented food as there are chances of mesophilic contaminants active in the fermented food if kept in exposure to contamination or the result of poor quality control while the packaging David Sturm has answered nearly the exact same answer. It can also be understood by a condition that while the food sample has been a fermented product and the time the other mesophilic contaminant was inactive due to certain reason and with due course of time after a certain time limit the microbe dominating the fermented food might start producing metabolites that other contaminant can consume. Although microbe in the food sample present in its packaging it can be slow but while being the mesophilic and getting the nutrient from the plate itself it has a good chances of experiencing a better growth then in the food itself.

Hope this might clear your doubt,

Best Regards

QuEChERS sample preparation

Also check https://www.tandfonline.com/doi/full/10.1080/21645698.2019.1649531

check this out

.... of most bacteria, including food pathogens, spoilage bacteria, and the lactic acid bacteria used in vegetable fermentations, are readily destroyed by heating to 160°F (71°C), especially when the pH is low. Acid and low pH are also toxic to most bacteria. https://www.ars.usda.gov/ARSUserFiles/60701000/FoodSafetyPublications/p328.pdf

According to Microbiology book by Mahon, Yersinia enterolitica it is mostly occur after ingestion of contaminated food, often pork and vacuum-packed deli meat, beef, lamb, chicken and possibly chocolate milk and water. It can also acquired from contact with household pets.

Implementation of environmental monitoring for Listeria monocytogenes should be based on risk assessment. There is no one program that fits for all. Looking for Listeria monocytogenes is of particular importance in the wet processing environment, which is conducive to Listeria growth. The monitoring program must look for the niches and harborage sites where there is a potential contamination of Listeria monocytogenes, this is not to be overlooked where RTE foods are exposed to the environment prior to to packaging and the packages foods are not subjected to a kill step. It is recommended to test product contact surfaces (for Listeria spp.) and occasionally testing finished product (for L. monocytogenes). This guidance might be helpful:

by PCR

Ok, let me try...

On Columbia blood agar (with defibrinated blood incorporated after sterilization of the base medium) Listeria monocytogenes will grow and will give also a clear β-haemolysis reaction.

On TSA growth of the bacterium is obvious, though not so abundant (i.e. small colourless colonies and slow growth on the agar plate). For this to occur, 0.6% yeast extract should be added, thus making TSAYE, which corresponds to the preffered general purpose medium for L. monocytogenes. In general, yeast extract seems to be an important ingredient in the media used for Lm, since its use is suggested also by ISO 11290 in blood agar.

On XLD Lm will not grow, I can tell you that. I came up with that mistake once...

For the other culture media you mention, I can't tell.

Regards!

Thankyou Jasim Al-Saadi and Carole C Tranchant for your kind expertise regarding the subject.

We are taking up a work on milk from peri-urban areas of Guwahati city. Although there are some literature on detection of M. bovisin milk samples, I am interested on application of molecular based techniques as well as other rapid methods. How far is it possible to detect M. bovis in milk by PCR? Other than PCR, is there any sensitive and specific rapid test for detection of M. bovisin milk samples?I shall be very happy to have the answers for food safety. 

Best way to identify mold/fungi accurately is by DNA analysis which can be relatively easy to perform using Polymerase Chain Reaction (PCR) techniques. The rRNA/rDNA sequences of fungal samples are commonly amplified using PCR and sent for DNA sequencing. I normally use universal primers such as the ITS1 and ITS4 for identification of fungal samples.

If you are not familiar with PCR techniques, the general guideline is:

1. Isolation of pure fungal sample on a suitable medium

2. Extraction of genomic DNA

3. Performing PCR amplification of sample using the chosen ITS primer

4. Sending PCR products for DNA sequencing

5. DNA sequence obtain can be analysed using online softwares such as BLAST by NCBI

You can refer to this publication for more info:

Hope this helps! All the best!

You'll need to use Raoults Law to predict the effect of solutes on water activity depression. The Aw calculation needs to include all solutes (I see that you have 0.5% salt, which will have some effect. Have used Propylene Glycol in food systems and managed to get down to 0.8 without too much problem.

I agree with dear Dr Debra Sharon Ferdinand-James

Dear Taghreed Hafez,

You may be able to produce water soluble chitosan by yourself!

Vahid

Nice

by LAMP vageli.

I think you can refer to FSSAI standards in India. They are available on the website of FSSAI (http://www.fssai.gov.in/home/food-standards/regulations/food-products-standards.html). You may find related product (often not exact what you look for).

All the best

Yes, you will need permits. Probably from both CDC and USDA/APHIS as these organisms are human and livestock pathogens. 

CDC: https://www.cdc.gov/phpr/ipp/

APHIS: https://www.aphis.usda.gov/aphis/ourfocus/animalhealth/animal-and-animal-product-import-information/ct_organisms_and%20_vectors/

Dear Razieh,

Please see the below given link.

Best Regards

Krishna

I am Hernan Ceballos and do conventional breeding in cassava. We have succeeded increasing carotenoids in cassava roots through conventional breeding. Although I could envision genetic transformation and gene editing competing technologies with my own area of expertise, I have always supported them. We need to solve the problems of millions of people by whatever means. It should not be a matter of competition between technologies, nor about regulatory issues. Dr. Potrykus team made a breakthrough way back with golden rice. It made the point of bioforitification on one hand, and the power of genetic transformation on the other. Yet about half a million children turn blind each year in Africa because of vitamin A deficiency. It is a shame for mankind that somebody, seating comfortably in an office (mostly) in Europe, thinks he/she is paying a service by preventing GOOD technologies reaching these children. The regulations related to the release of GMOs condemn them to blindness (and eventual death) and condemn me to eat tomatoes full of pesticides (as it is often the case in many developing countries). Regulation has taken away MY RIGHT to eat a Bt-tomato... So I totally support Dr. Portykus´statement about regulation. Hopefully, gene editing will relax things a bit and easy the way for common sense to prevail. Hernan

Do you express your protein content on dry matter basis or on product basis?

Longer baking may increase the dry matter content, so protein in dry matter is for me the best value.

Hi Sana,

The main problem is that neutrons interact with the atomic nucleus and convert an stable atom into a radioactive one. That is the basis for the nuclear fission employed in Nuclear Power Plants (for instance) where low energy neutrons (thermal neutrons) are captured by the U-235 nucleus with a subsequent fission giving rise to Ba-144 and Kr-90 and more neutrons (chain reaction). It means that the food could be converted into radioactive in case of neutron exposure. Ionising radiation with Co-60 (1.17 and 1.33 MeV), Cs-137 (1.5 MeV), electron beam (up to 10 MeV) and X-ray (up to 5 MeV) never will induce radioactivity in the food with such energies and, depending on the given dose (never higher than 10 kGy), the organoleptic properties will not be affected.

I hope it could help you.

Virgilio

What, precisely, is your aim? Do you just want to see the protein in the gel (e.g., for quantification), or do you want to purify the active enzyme (e.g., for kinetic measurements or industrial use)?

In the former case, I'd start with 7 M urea, 2 M thiourea, 10 mM DTT and 1%SDS. 

In the latter case detergents with fat-like structure (lyso-lipids, fatty acids, FOS-choline, LysoFos, neopentenyl glycols) may be able to to solubilise functional protein. In any case, I'd start with a careful literature search. A day in the library can save you months in the lab!

phytase It is an enzyme and passes several steps of purification ..Definitely  Aspergillus niger is GRAS 

Dear Sel,

Just to complete the answers you already received.You should to don't forget that winemakers are always trying to pick-up white grapes with a low acidity level (ph around 3,3) instead for red wines they  pick-up grapes in function of the total polyphenols amount and not only using acidity level as a key quality indicator. Even if the acidity still to be a key quality indicator. That is one reason why white wines have almost "always" lower pH than red wines. To sum up : white grapes pick up : aromas+acidity firstly ; to pick up red grapes : polyphenols concentration (anthocyanins+tanins) firstly.

Hope this helps.

Microbiology is not chemistry, therefore you can not expect "uniform" results when sampling raw foods. In the case of raw milk, particularly if it is not filtered and homogeneous (by industrial treatment or by integral sample preparation) results could differ. In particular fat content may change counts drastically, and fat tends to go at the top of your container. In addition depending on the hygiene of the industrial container (or machinery) from where you got the samples, there could be pieces of biofilm from the container or equipment walls. We do not know the containers or equipment from where you got the samples, and how actually you took and prepared them; but wild differences seems to point to the lack of uniformity of the milk and/ or the presence of biofilm particles (e.g. lack of hygiene).       

You can cook the meat so that all the bacteria are dead and ensure its sterile. Then go for doping with your Ecoli and use chemicals as you suggest as preservative.

I think another factor that has not been discussed yet is an indirect impact.  Crop failures/yield reductions will lead to crop price rises.  This will make affected crops more attractive targets for adulteration.  We have already observed these practices e.g. with spices being adulterated with nut derivatives leading to food safety issues. 

Dear Maliha,

Low molecular weight chitosan is very potent anti-fungal substance. The following references will help you to know the details:

From an analysis I conducted, I found no significant difference between levels of contamination among carcasses at different slaughter stages. However, other recommended hygiene indicators such as Total Aerobic Counts and Enterobacteriaceae significantly differed. Is it safe to conclude for now that Enterococci is not a reliable hygiene indicator for raw meat?

Hi Hoshi, thank you! The second article is close to what I'm looking for

When you perform the total plate count you will be able to observe the growth of fungus as well as bacteria.

Once there is growth on the plate you can perform different staining techniques and after microscopic examination you can perform confirmatory spore staining 

Lucinda - I would like to see the article you mentioned you were going to attach.  In the U.S. it has been well documented that antibiotics have been over prescribed, and still are.  Not completing a prescription is also a problem, and we should not forget that besides agriculture there is also some misuse for pets.

You can use pcr to detect the stx 1and 2 and other genes for attaching and effacing 

You won't get such direct reports. You must first do the basic characterisations like MW etcAnd then from any data bases for proteins you may get some data.

Are you concerned about the aflatoxin or the conjugate? As indicated above the aflatoxin itself is quite stable. The protein (BSA) and  (stability of)  the conjugate might be a different story.

You should look up standard analysis  documents/ standard such as British Pharmacopoeia    https://www.pharmacopoeia.com/  , some older version is downloadable  

the test is done by macro and microscopic methods as far as I know. the application is done in flour and cereals product as this type of food product is exposed to a such contamination due to bad storage or bad    hygiene  practice. However the attached file may help   

The decrease in pH may be due to production of organic acids. I just think the increase in pH observed after the decrease may be due to some of these acids (eg lactic acid) are metabolized (act as substrate) to support microbial growth. Because this reduces the concentration of the acid hence an increase in pH. To  ascertain this you need to measure titratable acidity (TA) too and see how that changes along the course of microbial growth.

Please, make sure the inoculation size  of bacteria and isolation  source must be from  Human sources 

This paper may be useful -

Yes the serotype of C. muytjensii strain 51329 is Cmuy O:2.  See

Jarvis, K.G., Yan, Q.Q., Grim, C.J., Power, K.A., Franco, A.A., Hu, L., Gopinath, G., Sathyamoorthy, V., Kotewicz, M.L., Kothary, M.H., Lee, C., Sadowski, J., Fanning,S., Tall, B.D., 2013. Identification and characterization of five new molecular serogroups of Cronobacter spp. Foodborne Path.Dis. 10, 343-352.

 

Excellent Stephane, Thank you so much. you have answered my question perfectly. 

For your information, I am developing a specific method for marine yeast isolation that is why there are yeast and mold together.

The number of yeast cells and mold is not known to me but both in the seawater sample.

I used the YPD broth as enrich media as  explained before for 2 days then used the culture to inoculate a fresh media for 2 days then inoculate a fresh media for another 2 days 

this method is published briefly here https://www.researchgate.net/publication/261719490_Marine_Yeast_Isolation_and_Industrial_Application

but now I am writing a research article with full details on the isolation method, characterization  and identification of marine yeast. 

so many thinks a gain and if you have supporting reference for what you explained will be very much appreciated 

Abdelrahman

Dear  Javier Amores.

Although the plates used are specific to S aureus, Staphylococcus can also grow other, but should be reported as unquantifiable or not found in the lowest dilution that did.

Annex a photo where you can see colonies of S aureus and other Staphylococcus. They are samples of cheese

You can look at these articles, it will help u. Good luck

Hi Suneel. I agree with the comments of Ng Seah Young. I guess that if you want to make a curd is because you are thinking in some kind of cheese with probiotics characteristics. L. casei Shirota's growth kinetics parameters are essential in this because some microorganisms are slow fermenters and are added in a belated stage of the process. As a starter you need a quick fermenter which acidifies the milk fast and coagulates the casein. My advice for you is to search for this data although is real what Jacob Simons says about Yakult. I think there are better options for what you want.

Dear Bruno, I recomend you M26-A, M07-A8 and M100_S21  NCCLS - CLSI protocols.

Hi,

The report published by Prof. Chris Elliott from Belfast Queen's University on the Horsegate scandal in the EU tackles adulteration, food fraud  and integrity in a lot of detail.

This is difficult to say, simply from these characteristics. If you have the possibility, prepare the 16S-RNA and sequence that. This is a highly conserved molecule, which will tell the taxonomy of your strain. Otherwise, go more systematically through a microbial identification protocol, which can be found in Bergeys Manual and other source.

Whether you treat an individual leaf as a replication or treatment, I suggest under all conditions you put one inoculated leaf in one bag/container for an unbiased sampling.

 As far as I know , the  bacterial cell densities in the human colon of 10¹¹-10¹² are the highest reported for any known ecosystem. In my studies, the level of spoilage bacteria have never exceeded 10¹⁰ CFU per gram of spoiled fish.

See the following article,

Roasting coffee beans (coffea arabica) artificially contaminated with ochratoxin A strongly reduces the analytical ochratoxin A content but not the genotoxic effects. Current Topics in Toxicology. Vol.9, 75 - 80, 2013

Before doing the PCR, you could separate prokaryotic from eukaryotic DNA  by the presence of methylated CpG motifs (usually not methylated in prokaryotic DNA, whereas the majority of them are methylated in eukaryotic DNA). Some column kits able to bind methylated CpG motifs does exist.

In Chile there is a regulation por "Aspergillus" (in general, i.e. all species) in BSL2

Without a doubt, OPA (o-phthalaldeyhyde) is the best method for the assesment of proteolytic activity in dairy produts. 

Church F. C., H. E. Swaisgood, D. H. Porter and G. L. Castinagni. 1983. Spectrophotometric assay using o-phaldialdehyde for determination of proteolysis in milk and isolated milk proteins. J. Dairy Sc. 66:1219-1227.

See attached paper. It appears that many more species can grow on PALCAM and ALOA

You'd better find the  precipitation or color of CS2 by chemical reaction, then you can detect it.

could you give more details on itis interaction?

I think if you are simply trying to apply a microscopic approach to look at your cultures then atomic force microscopy (AFM) might be a good choice, certainly over SEM since it can image sample under (near) physiological condition (under fluid, or maybe in the culture medium) and obtain images with similar or even better resolution than SEM. AFM sample preparation is relatively simple and it can prevent artifacts caused by some traditional EM sample prep techniques, which are BTW relatively harsh. For the experiment set-up you might want to start with image the two cultures separately to obtain information such as dimension and surface morphology of each cell line. You might even be able to discover some special surface features on each cell lines that can serve as marks in later experiments. Then you can image the co-cultured cells and look for the spectral organization. Of course there is no guarantee that this approach will work but you never know until you try it out.

BTW, for sample preparation, in the literature there are many studies using AFM to image E.coli, you may find some information that could help you in those papers.

Toxicity for using spices as a whole in cooking would be negligible - but if you talk of the active ingrediants separately and isolate them they are definately upto a certain extent - toxic - again the question is toxic against whom?

It has been reported that some strains of bacteria especially the Salmonella enter the viable but nonculturable state (VBNC) when they encounter environmental stresses, such as low temperatures, oligotrophic conditions, infiltration press, and biocides including heavy metals and ultraviolet radiation. VBNC bacteria are organisms that fail to grow and develop colonies on media, but their metabolic activity capabilities indicate that they are still alive. Add 3% Tween 20 or 1% catalase enabled cells to make it become culturable again, with resuscitation times of 48 h and 24 h, respectively.

It will depend on the size of your bacteriocin.  My guess is that most bacteriocins will NOT be concentrated in this manner as most of the ones from lactic acid bacteria (that have relevance as food pathogen inhibitors) are small molecules, around 2,000-3,500 mol. wgt, so unless they complex with some other molecules, you won't be concentrating them.  But that is OK, because you will be 'cleaning up' the sample from most of the larger molecules that will be retained, but I would suspect the bacteriocins would elute through the filter and be in the filtrate.  You can easily determine the amount retained/lost by doing activity titer before, and after Centricon ultrafiltration.  If you have a vacuum dryer, one easy way would be to use Sep-Pak C18 cartridges hooked up to a syringe.....pass the bacteriocin-containing culture broth through that....everything will bind to the C18 matrix (eluate should look clear; cartridge will appear 'brown' by retained media proteins), and then using step-wise 10-ml rinses pushed through the cartridge by way of a syringe....wash with Isopropanol:  0% (water), 10%, 20%, 30%, 40%, 50%, 60%, 100%......dry these down with a vacuum concentrator if available and resuspend with water/buffer of your choosing.  You will likely find that much of the 'brown' protein content elutes somewhere in the 10-20% fractions and hydrophobic bacteriocin molecules in the 30-50% fractions. A simple way to use hydrophobic nature of these molecules to your advantage in quasi-purification.  Similarly, another way would be ammonium sulfate fractionation....stirring different media samples with different levels of ammonium sulfate saturation to selectively precipitate bacteriocins (0% ammonium sulfate, 20%, 40%, 60%), centrifuge, decant, resuspend pellet with water/buffer and check to see which percent cutoff may selectively precipitate your bacteriocin peptide. If you produce your bacteriocin in MRS broth, ammonium sulfate will cause the Tween 80 to flocculate at the surface upon centrifugation and you may want to collect that as I often found that is where the bacteriocin is found....in that surface flocculated film that can actually be scooped off the top (and resuspend that in water).  Good luck....these procedures can be found in the literature.

I am not sure what you mean by 'synthetic phages'?  Do you mean, bacteriophages that are harvested manually from bacteria or recombinant DNA-based bacteriophages?  Presuming the former, the use of bacteriophages as a food preservative faces a difficult challenge in that the lytic bacteriophages that are generated on a particular host are subject to restriction modification by that host whereby the phage DNA is modified and not considered 'foreign' and not degraded. That is the main problem....the recovered bacteriophage work best on that one host strain, likely showing a very high efficiency of plaquing (EOP). The problem is, as a food preservative, the antimicrobial ( i.e., bacteriophage) should work equally well on all targeted strains, such as E. coli O157:H7 for instance.  If it is propagated on 1 strain of E. coli O157:H7, it should work on all E. coli O157:H7 strains and not just work well on the 1 strain it was propagated on. Otherwise it renders the preservative with little or no effect in practical commercial application.  Aside from that problem, is the problem with 'inhibition kinetics'....bacteriophage are relatively large particles (compared to acid chemical antimicrobial molecules like lactic acid), they are finite in number and therefore as they interact with the food matrix, they are 'tittered out of solution' so to speak and the amount remaining is that much less, and also they require a direct contact and injection of their DNA into the targeted host bacteria, so it would likely only have application on the surface of a food, not after it is mixed into a food (like ground beef) where the food matrix may bind and complex with bacteriophage particles, or coat/cover/conceal targeted bacteria. So even on the surface of a food, what kind of titer is required in solution to provide coverage of the surface with bacteriophage no further apart from each other than the length of a bacterial cell?....since once they land on a surface, they are not planktonic anymore and are somewhat fixed in position. Good luck.

Hi! Have you had a look at the paper in the link below? It might be helpful.

Good luck!

It is important to remember that we ingest thousand of bacteriophages every day with no harm to human health.  In fact, there is interesting evidence of symbiosis between animal and bacteriophages, ( http://www.nature.com/news/viruses-in-the-gut-protect-from-infection-1.13023) which makes sense if we consider we (humans) have evolve with the daily presence of bacteriophages.   

Bacteriophages are the most abundant organism in the biosphere, and FDA granted the GRAS (generally recognized as safe) status to those commercialized for use in food production

Hi Nikhil 

Its better to go for trichoderma sp., Pluerotus sp. along with dung slurry it enhances the decomposition rate and it takes  maximum of 3-4 months.

Thank you

Add some antioxidant and 2,4,5-T or heavy amount of 2,4-D

What precisely do you want to do? Hedonic or quantitative/descriptive sensory analysis? They demand totally different strategies. There are fairly good handbooks and even normalized methods for jury training for quantitative descriptive sensory analysis.

Regarding the organo-zeolites, Peter lego from UK has done some work in that direction.The followimh is the citation of the article.

The role of clinoptilolite in organo-zeolitic-soil systems used for

phytoremediation. Science of the Total Environment 363 (2006) 1–10

It may throw more light on this aspect. In india we are trying for organo-zeolites for enhanicng the use efficiency of organic manures. A report was presented at ASA symposium during 2012. 

I have reproduced the abstract below.

Zeolites Regulate Nitrogen Release From Manure-Amended Soil.

https://scisoc.confex.com/crops/2012am/webprogram/Paper75196.html 

See more from this Division: S08 Nutrient Management & Soil & Plant Analysis

See more from this Session: N Fertilizer Sources and N Use Efficiency: I

Tuesday, October 23, 2012: 9:05 AM

Duke Energy Convention Center, Room 211, Level 2

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Kulasekaran Ramesh, Indian Institute of Soil Science, Bhopal, India and Khandakar Islam, Soil, Water, and Bioenergy Resources, The Ohio State University, Piketon, OH

Manure-zeolite amendment of soil has recently been evolved as a novel approach to minimize reactive nitrogen formation and loss from agricultural fields. Zeolites are nanoporous secondary minerals having a high selectivity for cations especially ammonium ion which could be used for slow release of nutrients through organo-zeolite mixture concept. To study the effects of zeolite on regulating nitrogen release, a lab incubation study using dairy manure mixed with naturally mined zeolite (such as Clinoptilolite) was conducted at the Ohio State University South Centers at Piketon, USA during August and September of 2011. Zeolite and manure treatments were: Manure at 50-g oven-dried equivalent (M50); Zeolite + Manure at 1:5 ratio (ZM1:5), ZM1:10, ZM1:15, ZM1:20 and ZM1:30, respectively mixed with soil and replicated 6 times. The zeolite-manure amended soil in different treatment combinations were incubated at room temperature (~25 0C) for 7 days. The replicated samples of zeolite-manure treatments were taken every day over a period of 7 days and ammonium and nitrate concentration was measured calorimetrically. Results showed that the ZM1:15 had steadily and consistently release ammonium with a non-linear (parabolic) dynamics over time.

We may have collaborative work if possible. 

Hello, I think that there are many research about this area of interest.

Fusarium fujikuroi has not been found in any other species to the best of my knowledge. It has a closely related species F. proliferatum, which has many hosts from cultivated species. These two species have been found to interbreed and produce progeny. However, they have their specific hosts. Yes, Fusarium fujikuroi is pathogenic to rice and results in bakane disease.

Dear Krystian,

we are working on the influence of structure on crystallization processes. This is within the food area. So it would be interesting to check changes in structure. We need water soluble samples.

As I am partly in Warsaw, it could also make sense to meet some time.

Best regards,

Johannes

Food processing machinery encompasses a diverse category of equipment needed to process vegetables, meat products, marine goods and other food items. Totally integrated automation covers entire goods handling chain from reception to dispatch and integrates interfaces. Ethylene oxide/carbon dioxide mixture, which has a wide use in the food industry for treating processed and unprocessed foods. With considerably increased dosages it is also used for sterilizing food. It can also be used for sterilizing other materials.

I do believe it might involve "risk free business strategy"; however, it would depend upon, the regulatory agencies' capacity of intervention, as the business branding and as the trademark policies (protection). Certainly, any effort addressed to improve food safety and public health is always welcome, if sustained genuine' objectives.

Dear Ibrahim

fined in the attachment file the requested methodology