Cryptosporidium

Cryptosporidium

  • Frederick R. Masangkay added an answer:
    5
    What is the best method for isolation cryptosporidium oocysts from fecal samples?

    which method is better? Flotation or percipition?

    Frederick R. Masangkay

    Hi Marziye!

    Currently I, my colleague and research advisee students heve concluded isolation of Cyclo and Crypto in water samples using Glass-membrane filtration method and it worked wonders even if it is just a very cheap (but practically productive) method. we had high positivity of water samples from a local watershed.

    there are many literatures you will come about and the most common and cost effective is the flotation technique. there are different flotation techniques and from the results of the paper of my thesis advisee we can recommend as Sheather's flotation technique because it yields very good results, is so easy to prepare and contains no toxic/harmful substances.

    I highly recommend also a method by the CDC where you will make a fecal smear pass through a wire mesh in a cardboard hole about 3 to 5 mm in diameter. you can proceed to do modified acid fast staining immediately after drying and methanol fixing the smear. this is a modification of the Kato-Katz technique thick smear method.

    for details of the procedure please visit the CDC/WHO and look for Bench Aids for the diagnosis of intestinal Parasites (WHO 1994).

    as far as the recommended concentration of decolorizer for Modified Acid Fast Staining of Cyclo and Crypto Oocyst we used 2% sulfuric acid. our methods and results were verified by the Research Institute of Tropical Medicine Philippines and all steps in the methodology were the standards that they perform in identification of the same in fecal samples.

    I hope I was able to help you.

    best wishes Marziye!

  • Eljelani SALIM Salim added an answer:
    4
    Extracting Cryptosporidium oocysts directly from feacal?

    has any body tried using Qaigen stool mini kit for extracting Cryptosporidium oocysts directly from feacal. IIf so, what is the concentration you usually getting since I have tried using it and it gave me very lower concentration (2-10ng)?

    Eljelani SALIM Salim

    thanks Elha,

    I am appreciating your help. 

    Regards, 

    Eljelani 

  • Eljelani SALIM Salim added an answer:
    5
    Cryptosporidium DNA works first time after extraction but did not work for next time?

    Does anyone know why after extracting Cryptosporidium DNA from feacal samples, the first PCR works well but when I need to run another PCR from the same extracted DNA, it does not work on gel picture although the positive control was working. I am using bioline kit but this issue happened many times for me. I hope to find help from you. 

    Regards,

    Eljelani SALIM Salim

    THANKS ANDREW. Actually my concerns are not about the primers. The issue is that the PCR working well in the first attempt, but when I need to generate another products from the same DNA samples using the same primers, it dose not work. it seems surprising but that is what happend

  • Mohamed A El-Malky added an answer:
    7
    Do you know any available kit for DNA extraction from stool. I am working on cryptosporidium?

    Thanks in advance for your replies.

    Mohamed A El-Malky

    Thanks a lot Alexander for your kind reply

  • Eljelani SALIM Salim added an answer:
    7
    How can I concentrate Cryptosporidium oocyst from water sample?

    How to concentrate the Cryptosporidium oocyst? Is it same as to concentrate oocyst from wash solution of vegetable samples?

    Eljelani SALIM Salim

    thank you so much for replying.

    Regards,

  • Hassanain Al-Talib added an answer:
    7
    Any easy and effective method for extraction of Cryptosporidium ?

    Any easy and effective method for extraction of Cryptosporidium ?

    Hassanain Al-Talib

    Thank you Atif 

  • Pwaveno H. Bamaiyi added an answer:
    4
    Does anyone have a research protocol for multiplex PCR in Cryptosporidium spp?

    I want to find as many species of Cryptosporidum as may be present in cattle using faecal samples. Is there a way to do this using some sort of Multiplex PCR for all the different species that are known to infect animals?

    Pwaveno H. Bamaiyi

    Many thanks Dr Jesse Thomas, Dr Hugo Luca Abate and Dr Felippe Danyel for your kind response

  • David Bruce Conn added an answer:
    2
    Does anyone know the best Isolation method of Cryptosporidium oocysts in food?

    Does anyone know the best Isolation method of Cryptosporidium oocysts in food?

    David Bruce Conn

    Dear Md. Jiaur Rahman,

    This would vary considerably depending on what type of food you are dealing with.  The FDA-BAM chapter noted above would be one possibility for produce contaminated by fecal oocysts that can be rinsed off.  In other foods, such as marine shellfish (oysters, mussels, etc.) the oocysts may be internally sequestered and therefore not amenable to simple rinsing.  We have worked with several techniques over the years in both shellfish and produce, as well as fecal samples.  Again, it depends on your intended use.  Do you want to isolate these just for identification/diagnosis, or for use in experimental work?  Also, how accurate do you want to be?  Do you want to identify just any species of the genus Cryptosporidium, or do you want to rule out human-infectious, zoonotic, or non-human-infectious species.  The most accurate methods would include PCR, or fluorescent in-situ hibridization (FISH) multiplexed with immunofluorescent antibody methods.  I've listed some papers below, available on ResearchGate, which outline the latter method in shellfish sentinels, not only for Cryptosporidum, but also for Giardia and microsporidia. Other papers by Graczyk have used this method specifically on edible oysters.  The 2008 review paper by Graczyk and Conn summarizes some related ideas in reference to many groups of microbial pathogens.

    Good luck with your work!

    • Source
      [Show abstract] [Hide abstract]
      ABSTRACT: The capacity of synanthropic flies belonging to the families Calliphoridae, Sarcophagidae, and Muscidae to serve as mechanical vectors of Cryptosporidium and Giardia among livestock and wildlife in a multispecies agricultural complex in northwest Georgia (USA) was studied using fluorescent in situ hybridization (FISH) and immunofluorescent antibody (IFA) techniques. Flies from all three families were carrying viable Cryptosporidium oocysts and Giardia cysts internally and externally, and deposited them on trap surfaces. Furthermore, the contaminated flies and trap surfaces occurred in association with each of the four domestic animal units (beef, dairy, equine, and sheep), and in wildlife areas dominated by deer (Odocoileus virginianus) and Canada Geese (Branta canadensis). Cryptosporidium was isolated from 55.56% of the fly samples, whereas Giardia was isolated from only 7.94%. The highest numbers of Cryptosporidium were associated with cattle and wildlife areas, while Giardia occurred in greatest numbers in the sheep area. Cryptosporidium was isolated comparably from exoskeletal elutants and fly homogenates, but Giardia was isolated primarily from homogenates, indicating primarily internal transport in the latter. Surfaces visited by the flies (i.e., traps) became contaminated by both Cryptosporidium (18.89%) and Giardia (1.11%). In conclusion, these flies can serve as mechanical vectors of both Cryptosporidium and Giardia in all of these settings, and undoubtedly play a role in movement of these pathogens among the various host species.
      Full-text · Article · Feb 2007 · Vector Borne and Zoonotic Diseases

    + 2 more attachments

  • Eljelani SALIM Salim added an answer:
    4
    Has anyone tried using FTA cards for cryptosporidium DNA extraction? Is it possible subjecting fecal samples to FTA cards in other words?

    Have anyone tried using FTA cards for Cryptosporidium DNA extraction? in other words, is it possible subjecting fecal samples to FTA cards?

    i hope to find answer to this question since my research based on it.

    Reagards,

    Eljelani SALIM Salim

    are you subjecting the samples directely or after concentration (flotation or sedimentation), for example? because my DNA target is from Cryptosporidium oocysts.

    is there away of extraction DNA out of FTA desks?.

    I would be very grateful if you coul help me in this regard.

    Regards

  • Muhammed oda Mallah added an answer:
    24
    Can anyone help me in the diagnosis of Cryptosporidiun parvum in the attached pictures?

    This parasite stain with acid fast stain

    + 1 more attachment

    Muhammed oda Mallah

    Thank you for your attention my sister. best regards

  • Lawrence Broxmeyer, MD added an answer:
    6
    Where can I find age-structured incidence data for AIDS defining opportunistic infections starting from before AIDS epidemic in Sub-Saharan Africa?

    I am interesting in finding prevalence/incidence rates of specific opportunistic infections of young age groups (not adult) from before and after the start of the AIDS epidemic (~1980). 

    In particular I am looking for trends in: Tuberculosis, Cryptosporidium, and Non-Typhi Salmonella. 

    Does anyone know if such data exists and where I can find it? I would want to look in an area where HIV/AIDS is widespread, so somewhere in Sub-Saharan Africa would be ideal (or the whole region). 

    Lawrence Broxmeyer, MD

    "Where can I find age-structured incidence data for AIDS defining opportunistic infections starting from before AIDS epidemic in Sub-Saharan Africa?
    I am interesting in finding prevalence/incidence rates of specific opportunistic infections of young age groups (not adult) from before and after the start of the AIDS epidemic (~1980). 

    In particular I am looking for trends in: Tuberculosis, Cryptosporidium, and Non-Typhi Salmonella. "

    ________________________________

    Very well. But why go to WHO? Perhaps you should start by seeking statistical data and maps for Tuberculosis and M. avium, which still are the leading causes of infectious death in HIV/AIDS. Before such typical and atypical mycobacteria were proclaimed "AIDS-defining illness" by the likes of WHO, you will notice that they admitted that tubercular disease killed close to 3 million people a year. Then since it has magically  "defined" AIDS -  it is now purported that tubercular disease kills merely a million and a quarter annually. Interesting math.

  • Mosaab Omar added an answer:
    4
    How can I detect antigens of any parasites (for example Cryptosporidium) in faecal samples, and what is the solution or buffer?

    I want to ask you about something.

    if I want to detect antigens of any parasites (for example Cryptosporidium) in faecal samples.

    what is solution or buffer that I will used to extract parasite antigens?
    if possible, I need to know this solution with composition and formulation.

    Mosaab Omar

    Thank you very much for your valuable information, about PBS yes i can use, but i need a detailed list of the chemicals and buffers that i can use.

    Thank you again

  • David Bruce Conn added an answer:
    8
    What organisms and/or objects are most important as mechanical vectors of pathogens among livestock, wildlife, and potentially humans?

    Non-host mechanical vectors are frequently overlooked as transmitters and/or disseminators of pathogens in agricultural, sylvatic, and urban systems. This may be particularly important for zoonotic pathogens in agricultural ecosystems, but depending on the geographical location and the particular pathogens involved, there may be a range of possibilities.  For example, trachoma is transmitted among humans by synanthropic flies even in some urban settings.  We have been studying this at various locations, focusing on zoonotic enteric protists such as Cryptosporidium and Giardia, as well as microsporidia.  To gain a more comprehensive understanding of non-host vectors of pathogens around the world, I seek input on diverse specific experiences.

    David Bruce Conn

    Thanks for all the thoughtful responses/answers, which are very helpful to me.  I have attached here one of our papers on mechanical transmission by insect in a complex ecosystem where livestock, wildlife, and people occur in close proximity and interaction.

    • Source
      [Show abstract] [Hide abstract]
      ABSTRACT: The capacity of synanthropic flies belonging to the families Calliphoridae, Sarcophagidae, and Muscidae to serve as mechanical vectors of Cryptosporidium and Giardia among livestock and wildlife in a multispecies agricultural complex in northwest Georgia (USA) was studied using fluorescent in situ hybridization (FISH) and immunofluorescent antibody (IFA) techniques. Flies from all three families were carrying viable Cryptosporidium oocysts and Giardia cysts internally and externally, and deposited them on trap surfaces. Furthermore, the contaminated flies and trap surfaces occurred in association with each of the four domestic animal units (beef, dairy, equine, and sheep), and in wildlife areas dominated by deer (Odocoileus virginianus) and Canada Geese (Branta canadensis). Cryptosporidium was isolated from 55.56% of the fly samples, whereas Giardia was isolated from only 7.94%. The highest numbers of Cryptosporidium were associated with cattle and wildlife areas, while Giardia occurred in greatest numbers in the sheep area. Cryptosporidium was isolated comparably from exoskeletal elutants and fly homogenates, but Giardia was isolated primarily from homogenates, indicating primarily internal transport in the latter. Surfaces visited by the flies (i.e., traps) became contaminated by both Cryptosporidium (18.89%) and Giardia (1.11%). In conclusion, these flies can serve as mechanical vectors of both Cryptosporidium and Giardia in all of these settings, and undoubtedly play a role in movement of these pathogens among the various host species.
      Full-text · Article · Feb 2007 · Vector Borne and Zoonotic Diseases
  • Christopher Scocco added an answer:
    3
    Does anyone know good primers for Cryptosporidium meuitable fleagridis- for real-time PCR?

    I need good primers, that are 100% specific to Cryptosporidium meleagridis and suitable for real- time PCR reaction.

    And, possibly, also C meleagridis positiv DNA sample.

    Christopher Scocco

    Likewise, I have used PrimerQuest for the same thing and it gives you good results for primer design.

    http://www.idtdna.com/Primerquest/Home/Index 

  • Recep Tekin added an answer:
    1
    What drug can best be used to treat cryptosporidium infections?
    According to recent studies, which drug is more effective: azitromycin, paromomycin, nitazoxanide, or spiramycin?
    Recep Tekin
    specific treatment:

    Nitazoxanide shortens duration and reduces mortality in malnourished children.[ Nitazoxanide is well tolerated with a good safety profile.
    Nitazoxanide, paromomycin and azithromycin are only partially effective and results with cryptosporidiosis in AIDS patients remain disappointing.
    Trials are small and evidence is conflicting. Drugs include the aminoglycoside paromomycin,and macrolides such as azithromycin and clarithromycin.
  • Venu Ravipati added an answer:
    1
    How many days can a Cryptosporidium positive faecal sample can be stored in --20C with out any preservative?
    Faecal sample storage without any preservative
    Venu Ravipati
    My observation is not more than a month in -20C without preservation; but can be extended with 2.5% Potassium dichromate solution for another 3-4 months at _20C
  • Michelo Simuyandi asked a question:
    Open
    Where can I get Cryptosporidium antigen?
    I am working on detecting Cryptosporidium specific antibodies in serum using ELISA and am looking for Cryptosporidium antigen (both C. Parvum and C. Hominis).
  • Tatiana taty Charles added an answer:
    3
    Other than those pathologies, what severe pathology could cause the cryptosporidium?
    I read your interesting article and I see that you especially mention severe pathologies such as diarrhea in case of immunodeficiency and malnutrition.
    Tatiana taty Charles
    I thank you Dr for your answer. Pancreatitis is also a common severe pathology encountered in cryptosporidium infection. But i don't find in literature if it can happen late or spontaneously after infection
  • Ma. Del Socorro Carreno added an answer:
    9
    What is the best (validated) disinfectant for Cryptosporidium in clinical lab?
    What is the most ideal (validated) disinfectant for Cryptosporidium in clinical lab?
    Ma. Del Socorro Carreno
    Hi, I find, this information:
    Cryptosporidium have a spore stage (oocyst) and in this state for long periods can survive outside the host and can also withstand many common disinfectants, especially based disinfectants cloro.3 Due to this resistance, water treating to remove Cryptosporidium generally based on coagulation followed by filtration or boiled. In 2007 it was discovered that Cryptosporidium is sensitive to ultraviolet light and ozonation, May 4 having already installed such water treatment systems in cities where they have had problems with the presence of parásito.6

    Most treatment plants that take water from rivers, lakes and reservoirs for public drinking water production use conventional filtration technologies. This involves a series of processes including coagulation, flocculation, sedimentation and filtration. Direct filtration, which is generally used for treatment of water with low levels of particles includes coagulation and filtration, but no sedimentation. Other common filtration processes are sand filters or diatomaceous earth, membranes and bag and cartridge filters.

    Conventional technologies direct sand and diatomaceous earth can remove 99% of Cryptosporidium.7 membranes and the bag and cartridge filters can remove Cryptosporidium on the basis of a product due concreto.Con concentration and contact time , can be performed Cryptosporidium inactivation treatment with chlorine dioxide and ozone. Can also be performed with ultraviolet light treatment relatively low doses.
    3.↑ «Chlorine Disinfection of Recreational Water for Cryptosporidium parvum». CDC. Consultado el 06-05-2007.
    4.↑ «Irreversible UV inactivation of Cryptosporidium spp. despite the presence of UV repair genes.». NCBI PubMed. Consultado el 06-05-2007.
    5.↑ «Ultraviolet Disinfection and Treatment». AWWARF.org. Consultado el 06-05-2007.
    6.↑ «Östersund to flush parasite from water pipes.». Radio Sweden. Consultado el 25-04-2011.
    7.↑ «The Interim Enhanced Surface Water Treatment Rule – What Does it Mean to You?» (pdf). USEPA. Consultado el 06-05-2007.

About Cryptosporidium

A genus of coccidian parasites of the family CRYPTOSPORIDIIDAE, found in the intestinal epithelium of many vertebrates including humans.

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