Diagnostic Parasitology FAQs (Garcia)
Splitting the stalks and crowns of these plants may reveal discoloration and decay which can limit water movement up the plant, leading to decline and death of the plant and its leaves. A minute staining time appears to work better than 15 min; staining times will depend on stain dilution. Page Macintosh printing Click Print. It is anticipated that two more species will be added as well as more ingredients, including some branded products. Iowa State University Economist Advises Cost Containment, and Patience, for Farmers Marketing Crop As harvest approaches, Chad Hart, associate professor of economics at Iowa State University, urged Iowa farmers to watch their costs and be patient as they face a difficult market for corn and soybeans. You need to set up the network protocols on the machine to use it as your network machine.
Here is the collection of some of the Verses in Quran about Ramadan. It is one of the last ten odd nights in the month of Ramadan and is full of blessings. This month of Ibadah ends with the Muslim festival of Eid ul Fitr. Learn How to determine the Date of Lailatul Qadr.
Tarawih are the extra prayers some Muslim Communities perform at night after Isha Prayers in the Islamic month of Ramadan. They are not mandatory Prayer but are still of utmost Importance. Zakat is another Pillar of Islam, and giving Charity becomes even more important during Ramadan.
It is a way to purify your wealth for the will of Allah SWT and is payable on assets owned over one lunar year. The collected Zakat is required to be given to the poor and deserving people.
In Ramadan, all good deeds are rewarded more than in any other month of the year. This is the why many people choose give Zakat Sadqa to poor in this Month. Here are the Ways to be charitable in Ramadan.
Itikaf means to be in isolation in a Masjid or at home with the intention of solely dedicating your time to the worship of Allah SWT. It is Sunnat-al-Muaqidah Sunnah that is urged to be performed to sit in Itikaf in the last 10 days of Ramadan. A person may commence Itikaf after sunset of 20th of Ramadan, and end it when the moon for Eid is sighted.
The Sunnah stays the same if the month of Ramadan is of 29 or 30 days. Here is All you need to know about Itikaf in Ramazan. Search for a City or Zip to set your location. Disclaimer All information on IslamicFinder. Wednesday, May 16, See all Islamic days. Q What is Ramadan? A Ramadan is the 9th month in the Islamic Calendar.
Muslims all over the world fast during this month. A Ramadan is expected to start on the 16th of May , Wednesday. The date is tentative and may change subject to sighting of the moon. A Ramadan will end on Thursday, 14th of June A Ramadan is considered to be one of the most blessed months.
Ramadan brings about an air of festivity for the Muslims. The whole month is celebrated to thank Allah for the blessings that have been bestowed upon us and to learn self-control. A The main purpose of Ramadan is to teach Muslims self-control and perseverance.
The BD MAX System automates sample lysis, nucleic acid extraction and concentration, reagent rehydration, nucleic acid amplification and detection of the target nucleic acid sequence using real-time polymerase chain reaction PCR.
Amplified targets are detected with hydrolysis probes labeled with quenched fluorophores. Gloves must be changed before manipulating reagents and cartridges. Do not use expired components. It indicates the presence of DNA from E. As with all PCR-based in vitro diagnostic tests, extremely low levels of target below the analytical sensitivity of the assay may be detected, but results may not be reproducible. False negatives may also occur if the level of organism nucleic acid is below the threshold of detection.
False negative results may occur due to loss of nucleic acid from inadequate collection, transport or storage, or due to inadequate organism lysis. The SPC has been added to the test to aid in the identification of specimens that contain inhibitors to PCR amplification.
The SPC does not indicate if nucleic acid has been lost due to inadequate collection, transport or storage, or whether cells have been inadequately lysed. BD MAX Enteric Parasite Panel results may or may not be affected by concurrent antimicrobial therapy, which may reduce the amount of target present.
The performance of this test has not been established for monitoring treatment of Giardia lamblia , Cryptosporidium hominis and C. This test is a qualitative test and does not provide quantitative values nor indicate the quantity of organisms present. The performance of this test has not been evaluated for immunocompromised individuals or for patients without symptoms of gastrointestinal infection.
The effect of interfering substances has only been evaluated for those listed in this labeling. This test is a qualitative test and does not provide a quantitative value for the organism s in the sample. The FilmArray GI pouch is a closed system disposable that houses all the chemistry required to isolate, amplify and detect nucleic acid from multiple gastrointestinal pathogens within a single stool specimen.
The rigid plastic component fitment of the FilmArray GI pouch contains reagents in freeze-dried form. The flexible plastic portion of the pouch is divided into discrete segments blisters where the required chemical processes are carried out.
All other operations are automated. Two hundred microliters is the sample volume requirement. The FilmArray GI Panel test consists of automated nucleic acid extraction, reverse transcription, amplification, and analysis, with results available in one hour per run i.
FilmArray GI Panel runs were considered valid if the run completed normally and internal controls passed. Stool samples may contain a high concentration of organisms. To avoid possible contamination, samples should be processed in a biosafety cabinet. If a biosafety cabinet is not used, a dead air box, a splash shield, or a face shield should be used when preparing samples.
A biosafety cabinet or work station that is used for performing stool pathogen testing e. To avoid residue build-up and potential PCR inhibition, wipe disinfected surfaces with water. Samples and pouches should be handled one at a time. Change gloves and clean the work area between each sample.
Specimens should be processed and tested as soon as possible, though they may be stored at room temperature or under refrigeration for up to four days. The freeze-dried yeast is present in the pouch and becomes rehydrated when the sample is loaded. A positive control result indicates that all steps carried out in the FilmArray GI pouch were successful.
A positive result indicates that 2nd stage PCR was successful. Good laboratory practice recommends running external positive and negative controls regularly. Enteric transport media can be used as an external negative control.
Previously characterized positive stool samples or negative samples spiked with well characterized organisms can be used as external positive controls. External controls should be used in accordance with the appropriate accrediting organization requirements, as applicable. Once melt curves have been identified, the software evaluates the three replicates for each assay to determine the assay result.
Assays that do not meet these criteria are called negative. If four or more distinct organisms are detected in a specimen, retesting is recommended to confirm poymicrobial result.
Detection of approximately 23 different Cryptosporidium occurs, including the most common species of human clinical relevance i. The assays do not differentiate between Cryptosporidium spp. A positive result for either or both assays will give a Cryptosporidium Detected test result. This assay may cross-react with the closely related E. The Result Summary section of the test report lists the result for each target tested by the panel. See Results Summary section in the Instruction Booklet for detailed information about interpretation of test results and appropriate follow-up for Invalid results.
It has not been validated for use with other stool transport media, raw stool, rectal swabs, endoscopy stool aspirates, or vomitus. Test performance has not been established for patients without signs and symptoms of gastrointestinal illness. Virus, bacteria, and parasite nucleic acid may persist in vivo independently of organism viability.
Additionally, some organisms may be carried asymptomatically. Detection of organism targets does not imply that the corresponding organisms are infectious or are the causative agents for clinical symptoms. The performance of this test has not been established for monitoring treatment of infection with any of the panel organisms. Austin, TX [14, ]. The xTAG GPP is a multiplexed nucleic acid test for the simultaneous qualitative detection and identification of multiple viral, parasitic, and bacterial nucleic acids in human stool specimens from individuals with signs and symptoms of infectious colitis or gastroenteritis.
The target is amplified using polymerase chain reaction PCR or reverse transcription PCR, then analyzed with Luminex xTAG technology to detect the presence or absence of each pathogen in the panel. The test detects 11 gastrointestinal pathogen targets: Additionally, simultaneous molecular testing on a single sample within a single shift provides significant benefit to laboratories in terms of workflow and resource utilization. Human stool samples are pretreated and then subjected to nucleic acid extraction.
A signal or median fluorescence intensity MFI is generated for each bead population. A single multiplex reaction identifies all targets. All results should be used and interpreted in the context of a full clinical evaluation as an aid in the diagnosis of gastrointestinal infection. Confirmed positive results do not rule out co-infection with other organisms that are not detected by this test, and may not be the sole or definitive cause of patient illness.
Analyte targets virus, bacteria or parasite nucleic acid sequences may persist in vivo, independent of virus, bacteria or parasite viability. Detection of analyte target s does not guarantee that the corresponding live organism s is present, or that the corresponding organism s is the causative agent for clinical symptoms.
Positive and negative predictive values are highly dependent on prevalence. False negative test results are more likely prevalent when disease is high. False positive test results are more likely during periods when prevalence is low. This test is a qualitative test and does not provide the quantitative value of detected organism present.
Known strains or positive clinical samples for the targeted viruses, bacteria or parasites should be included in routine quality control procedures "external controls" as positive controls for the assay. One or more of these external controls are analyte positive controls and should be included with each batch of patient specimens. Controls positive for different targets should be rotated from batch to batch. External controls should be prepared, extracted and tested in the same manner as patient samples http: Failure to correctly interpret test results in the context of other clinical and laboratory findings may lead to inappropriate or delayed treatment.
For example, a microorganism present as a colonizer may be correctly detected, but not be the true cause of illness. Although this identical risk would be present from use of any microbiological assay in this setting, simultaneous testing of multiple analytes in a multiplex assay may be more likely to detect an unanticipated colonizer that might not be tested for individually.
Both clinical presentation and other results would likely substantially mitigate concerns with both false positive and false negative test results. There are also several molecular tests that are in clinical trials for the detection of select gastrointestinal parasites.
These tests are molecular gastrointestinal panels and target the most commonly occurring parasitic stool pathogens. Although there are laboratory developed tests for most parasites, these are not commercially available or available only in specialized testing centers.
When such tests are used, there should be attention given to the use of internal amplification controls to detect inhibition, since common specimens, such as blood and stool, contain PCR inhibitors. Thorough validation is required before these are implemented for clinical testing. Specific organisms under consideration include: Entamoeba histolytica, Blastocystis hominis Blastocystis spp.
Blood parasites include Plasmodium spp. What is the most effective technique for the identification of the intestinal protozoa? Although some protozoan cysts can be seen and identified on the wet preparation smear direct mount, concentration sediment wet mount , the permanent stained smear is the procedure that is recommended as the most relevant and accurate for the identification of the intestinal protozoa.
These preparations are examined using the oil immersion objective X for a total magnification of X At least oil immersion fields should be examined prior to reporting the permanent stained smear result. Are trophozoites ever seen in the wet mounts of stool? Usually the trophozoites are not seen in the concentration sediment wet mount preparations unless they are prepared from stool preserved in Universal Fixatives.
Motile trophozoites can occasionally be seen in the direct wet smear, but the number of times this occurs is rare.
What are some of the tips to consider when reporting Entamoeba hartmanni? When you see a cyst on the permanent stained smear, there is often a halo representing shrinkage. The cyst needs to be measured to include that halo. On the bench, the measurements for this cyst generally run from around 9. Also, on the bench, the E. Do nonpathogenic protozoa ever cause symptoms? Endolimax nana, Iodamoeba bütschlii, Chilomastix mesnili, and Pentatrichomonas hominis as examples have been categorized as nonpathogens.
Although rare, patients have been documented who have been symptomatic with a nonpathogen. However, it is sometimes difficult to determine from the case history the extent of the workup, including coccidia and the microsporidia. Before assigning symptoms to nonpathogenic protozoa, a comprehensive search for other proven pathogens should be performed.
One can also report the true pathogen if the fecal immunoassay specific for Entamoeba histolytica is positive. What color is the autofluorescence seen with Cyclospora cayetanensis?
The color depends on the particular FA filters used. If the Calcofluor white filters are used, the oocysts will appear as pale blue rings; if the FITC filters are used, the oocysts will appear to be more yellow-green. Why is Blastocystis hominis so controversial regarding pathogenicity? What we currently call Blastocystis hominis morphologically appears to be approximately 10 different strains or species, some of which are pathogenic and some are nonpathogenic.
Based on continuing molecular studies, changes in the classification of this group are probable. Unfortunately, all of the strains or species are morphologically identical; therefore, the correct identification reported should remain Blastocystis hominis , and the number should be quantitated from the permanent stained smear rare, few, moderate, many, packed.
Some laboratories have begun reporting Blastocystis spp. This is the recommended approach. Some helminth eggs are quite heavy unfertilized Ascaris eggs and will not float, even using zinc sulfate with a specific gravity of 1. Thus, both the surface film and the sediment should be examined before reporting the specimen as negative. It is important to make sure that the larvae seen are, in fact, the rhabditiform non-infectious larvae of S.
The agar plate culture is the most sensitive method for the recovery of S. Also, remember that migrating nematode larvae could also be recovered from respiratory specimens sputum, BAL, etc. Are there any specific recommendations for the detection of schistosome eggs? When trying to diagnose schistosomiasis, regardless of the species suspected you should be examining both stool several different stool specimens and urine spot urines plus 24 hour specimen collected with no preservatives.
Occasionally adult worms get into blood vessels where they aren't normally found Example: When you perform a sedimentation concentration, use saline to prevent premature egg hatching. Once you are ready to try a hatching procedure, then you can put the sediment into spring water dechlorinated water to stimulate hatching.
When examining wet mounts under the microscope, you need to look for the movement of cilia on the miracidium larvae within the egg shell thus the need to collect the specimens with no preservatives. You want to be able to tell the physician whether the eggs, if present, are viable or all you see are dead egg shells. If you suspect schistosomiasis, it is recommended that you examine a number of wet mounts, particularly if you aren't going to perform a hatching test.
What is the most sensitive test for the diagnosis of strongyloidiasis? Agar plate cultures are recommended for the recovery of S. Stool is placed onto agar plates and the plates are sealed to prevent accidental infections and held for two days at room temperature. As the larvae crawl over the agar, they carry bacteria with them, thus creating visible tracks over the agar. In a study looking at the prevalence of S. These results indicate that the agar plate approach is probably a much more sensitive diagnostic method and is recommended for the diagnosis of strongyloidiasis.
It is important to remember that more than half of S. The agar plate method continues to be documented as a more sensitive method that the usual direct smear or fecal concentration methods. Daily search for furrows on agar plates for up to 6 consecutive days results in increased sensitivity for diagnosis of both S.
Also, a careful search for S. Serologic testing for antibody is also recommended if strongyloidiasis is still suspected in the absence of laboratory confirmation of larvae.
Where can one get serologies for a Baylisascaris procyonis infection? CDC M-F 7: Should common or scientific names be used when reporting the presence of parasites? What happens if there are several different names used in the literature Giardia lamblia, G. It is appropriate to use the most commonly accepted name Giardia lamblia ; you can also let the proficiency testing organism lists be your guide. Remember, it is very important to notify all clients prior to making any name changes.
How has the reporting of Cryptosporidium parvum changed and why? It is now known that more than one species of Cryptosporidium can cause disease in humans C.
However, the different species cannot be differentiated on the basis of morphology. It is now recommended that the more correct reporting format would be: The reporting and quantitation rare, few, moderate, many of WBCs PMNs, macrophages, eosinophils provides some additional information for the physician.
If the patient continues to have diarrhea, it may give the physician something more to consider, particularly if stool culture has not been ordered. Also, conditions related to non-infectious diarrheas may also result in WBCs and macrophages in the stool. Physicians can then decide on the relevance of the information. Remember, that when reporting WBCs, we can also identify eosinophils from the permanent stained smear may or may not be related to parasitic infection and may or may not correlate with peripheral eosinophilia — this information may also be helpful.
Reporting yeast is a bit different. In order to report anything about yeast, you must know the stool was fresh or immediately put in preservative. However, if you don't know whether the collection criteria were met, then reporting anything about yeast is NOT recommended since this type of report may be misleading. How should Blastocystis hominis be reported? In the past there was some agreement that the number of organisms present mod, many, packed were more likely to be associated with symptoms.
However, in the past few years, there have been anecdotal reports of patients being symptomatic with rare or few organisms, as well. The current recommendation is to report the organism and quantitate per rare, few, mod, many, packed using your proficiency testing quantitation scheme.
It is important to confirm that the physicians know what the report means and understand the controversial issues surrounding this organism - they can then correlate the numbers with symptoms. If rare or few, there is no solid data per numbers relevance other than the anecdotal case reports many of which are word of mouth.
Based on continuing molecular studies, changes in the classification of this group could be updated. Unfortunately, strain or species differences cannot be detected by morphology; they look alike. Some labs are now reporting Blastocystis spp. You should begin reporting Blastocystis spp. It is also recommended that a report comment be included regarding strains, subtypes, or species and pathogenicity.
How should intestinal protozoa be reported? This has been the accepted way of reporting intestinal protozoa for a couple of reasons: Different drugs are used to treat Entamoeba histolytica cysts vs. Also, since the cyst form is the infective stage for the protozoa exception: For these reasons, the current recommendation is to continue to report all protozoa genus, species, and stage.
Isospora belli oocysts coccidia can be detected from the concentration sediment examination. The report of Blastocystis hominis should be accompanied by the following report comment: This explains why some patients are symptomatic and some are asymptomatic.
Trichuris trichiura eggs, Clonorchis sinensis eggs, schistosome eggs also report viability of Schistosoma spp. Charcot-Leyden CL crystals 2. Why are all proficiency testing specimen answers reported and quantitated, while clinical specimen reports are rarely quantitated in terms of organism numbers? There are two issues to consider.
Some exceptions might be: Trichuris trichiura eggs in a concentration wet mount light infections might not be treated. Since many organisms are shed on a random basis, quantitation may change dramatically from day to day and generally has little clinical relevance. How should you examine a wet preparation for proficiency testing? Most directions will recommend you shake the vial and take a sample from the mixed vial contents.
However, if the number of eggs is few, you may be better off taking a very small drop from the settled material. If you take too large a drop, it will be too thick to examine properly.
Make sure you do not add too much iodine; very darkly stained helminth eggs can resemble debris. The microscope should be aligned properly using Köhler illumination. Make sure the light is not too bright; otherwise, you may shine right through the organisms and miss the parasites. Also, you may want to close the diaphragm a bit to provide a bit more contrast. Extra contrast is particularly important when reading with saline only no iodine added.
How should you examine a permanent stained fecal smear for proficiency testing? Make sure the light is very bright condenser all the way up and the diaphragm is open.
Review both thin and thick areas of the smear; examine at least oil immersion fields using the X oil immersion objective before you report the specimen as a negative. If you use the 50X or 60X scanning oil immersion lens, you still need to review oil fields as indicated above using the X oil immersion objective.
How should tissues be submitted to the laboratory? Tissue specimens should be immediately sent to the laboratory and kept moist during transit. If the tissue will be processed for culture Acanthamoeba, Naegleria, Leishmania spp. If the tissue will be processed for wet examinations and permanent stained smears, any type of container is acceptable. Remember that the specimen must be kept moist; if the specimen dries out during transit, neither culture nor stained smears will provide acceptable results.
How should eye specimens be submitted for Acanthamoeba culture? This will prevent the small tissue specimen from drying out if during transit it is shaken onto the side of the container.
If the container is not full of fluid, the tissue may dry out and will be unacceptable for culture. How should duodenal drainage specimens be handled?
Duodenal drainage specimens must be submitted to the laboratory as quickly as possible for processing. Delays may prevent organism recovery and identification. These specimens may be very liquid and may contain a lot of mucus. Many questions have been asked regarding various aspects of diagnostic medical parasitology and the examination of blood specimens. Specimen Collection Blood 1. If Plasmodium falciparum parasites are sequestered in the capillaries, why not do a finger stick, rather than a venipuncture?
The capillaries are generally in the deep tissues spleen, liver, bone marrow , so a finger stick blood sample is no more likely to be positive than a venipuncture blood. There may be some differences with P. What is the best anticoagulant to use for blood specimens? Also, when using anticoagulants, the proper ratio between blood and anticoagulant is necessary for good organism morphology.
Heparin can also be used, but EDTA is preferred. Finger stick blood is recommended when the volume of blood required is minimal no other hematologic procedures have been ordered. However, finger stick blood is no longer commonly used in many parts of the world.
When blood is collected in EDTA, specimens should be processed immediately after blood collection. Parasite numbers may decrease if processing is delayed, even 4 to 6 hours. Adhesion of the blood to the slide can be a problem if the ratio of anticoagulant to blood is high, the patient is anemic, or the blood was held in EDTA too long.
Why do new slides have to be precleaned with alcohol prior to use? When should blood specimens be drawn for a suspect malaria diagnosis? The majority of patients we see in the US with malaria have never been exposed to the organism before, thus they have no antibody. These immunologically naïve patients may present with nonspecific symptoms that can mimic many other diseases. The rule of thumb is to draw immediately ; do not wait for some "magic" periodic cycle that may never appear.
Patients with a very low parasitemia with P. With any patient where malaria is suspect or the patient has a FUO fever of unknown origin , blood should be drawn and both thick and thin blood films prepared and examined immediately. This request is always considered a STAT request.
In endemic areas of the world where people have been exposed to the parasite before have some antibody , they may not become symptomatic until they actually have some sort of periodic fever cycle. However, you should always use the general guideline to "draw" immediately. Also, remember that one set of negative blood films does not rule out malaria.
If the first set both thick and thin films is negative, you can recommend that additional blood films be drawn in about hours. Also, any decision to delay treatment should be left to the physician, not the laboratory. Why is it important that the EDTA blood be processed as quickly as possible?
If a tube of blood containing EDTA cools to room temperature and the cap has been removed, several parasite changes can occur. The parasites within the RBCs will respond as if they were now in the mosquito after being taken in with a blood meal.
The morphology of these changes in the life cycle and within the RBCs can cause confusion when examining blood films prepared from this blood. Why is it important to keep thick films from getting hot heat fixation? Heat will fix the RBCs and they subsequently will not lyse in the staining process.
See 1, E above. The laked thick films and thin films should be dipped in absolute methanol and placed in a vertical position to air dry. This is particularly important if the blood films will be stored for days or weeks prior to staining. Processing of patient specimens for diagnosis needs to be performed on a STAT basis; it is inappropriate to hold slides for staining unless they are for teaching purposes and time is not a critical issue. However, the longer the slides are held, the more likely there will be a decrease in overall staining quality.
Why is it important to always examine both thick and thin blood films prior to reporting the specimen as negative for blood parasites? Thick blood films allow a larger amount of blood to be examined, which increases the possibility of detecting light infections.
However, only experienced workers can usually make species identification by thick film, particularly for malaria parasites.
The morphologic characteristics of blood parasites and identification to the species level are best accomplished from the thin films. However, it is mandatory that both types of films be examined prior to reporting negative findings. What are the advantages of the thick blood film?
The aims of the preparation of a thick film is to have a drop blood with 20 or 30 layers of red bloods on the slide, then to lyse the red cells, wash off the hemoglobin and stain the parasites, which remain intact in the process.
As a consequence, red blood cells will not normally be visible, but white blood cells and parasites will be seen. It is essential to get used to the characteristics of malaria parasites and know what to look for on such a preparation. An experienced microscopist should be able to detect 20 parasites per microliter of blood i.
A greater volume of blood can be examined in the same amount of time taken for examination of the thin blood film. The presence of phagocytized malaria pigment within WBCs particularly in cases with low levels of parasitemia can be very helpful. What are the disadvantages of the thick blood film? Recognition of organism distortion and ID to species are generally more difficult from the thick film. What are some of the problems associated with thick blood films? The thick film flakes off during the staining process.
What are some of the tips for improving thick blood films? After the thick films are dry, fix the smear with acetone for a few seconds ONLY dip twice , then air dry the film. These films tend to have a clean background, making the parasites easier to see.
Overall adherence to the slide is enhanced using this approach. How do you prepare thin blood films? Before fixing the thin blood films in absolute methanol, the film must be completely dry. If slides must be stored unfixed, they should be frozen. It is better to use dispensing bottle for methanol, rather than a Coplin jar, in which the methanol will pick up water from the air.
Methanol used on one day Coplin jar should not be reused the next day — begin with fresh stock. What are the advantages of the thin blood film? It is much easier to identify malaria organisms to the species level from the thin blood film. The parasitemia can be calculated much easier from the thin blood film than the thick film.
What are the disadvantages of the thin blood film? This method has a much lower sensitivity than the thick blood film; thus, infections with a low parasitemia may be missed. What are some of the problems associated with thin blood films? Poorly prepared thick and thin blood films can be seen in Figure 2.
Films are too thick. The combination thick-thin blood film provides both options on one glass slide and the slide can be stained as either a thick or thin blood film Figure 3. If fixed prior to staining, then the smear will be read as a thin blood film; if RBCs are lysed during staining, the preparation will be read as a thick blood film parasites, platelets, WBCs.
This combination blood film dries more rapidly than the traditional thick blood film, thus allowing staining and examination to proceed with very little waiting time for the slide s to dry.
How should malaria smears be stained with Giemsa Stain? Giemsa is a mixture of eosin and methylene blue. Stock solutions of Giemsa may be purchased commercially. Some brands are better than others. The stock solution of Giemsa stain is easily prepared from commercially available Giemsa powder.
Although most people do not filter the working stain prior to use if using a Coplin jar , results are better overall if the working stain is filtered through Whatman no. Make sure to use absolute methanol acetone-free for fixing thin blood films. Stock buffered water is stable for 1 year at room temperature. Working stock buffered water is stable for 1 month at room temperature. Stock Giemsa stain is stable at room temperature indefinitely; stock stain appears to improve with age similar to that seen with iron-hematoxylin stains.
A minute staining time appears to work better than 15 min; staining times will depend on stain dilution. How should you handle a delay between thick film preparation and staining?
Thick films can be preserved, particularly if there will be a delay prior to staining. Dip the slides in a buffered methylene blue solution 0. Can blood stains other than Giemsa stain be used to stain the blood films? The detection of parasites in the blood has been made possible by the use of Romanovsky-type differential stains which selectively color the nuclear material red and the cytoplasm blue.
This reaction takes place under optimal pH conditions: It is more appropriate to use a stain with which you are familiar, rather than Giemsa which is somewhat more complicated to use. PMNs will serve as the QC organism for any of the blood stains. Any parasites present will stain like the PMNs, regardless of the stain used. Also, the CAP checklist does not mandate the use of Giemsa stain.
How should malaria blood films both thick and thin films be examined? A minimum of oil immersion fields using the X objective should be examined. The blood film can be scanned using a 50X or 60X oil immersion lens, but final reporting of the results should be based on the use of the X oil immersion lens for a total magnification of X1, What type of QC slides should be used for blood parasite work?
What is a good source for teaching slides? A good source for teaching slides is: They have slides and specimens available for sale. Contact them for a brochure: Schizogony occurs in the internal organs spleen, liver, bone marrow, etc. Ischemia caused by the plugging of vessels within these organs by masses of parasitized RBCs will produce various symptoms, depending on the organ involved. Onset of a P. The onset is characterized by fever, a more severe headache, and nausea and vomiting, with occasional severe epigastric pain.
There may be only a feeling of chilliness at the onset of fever. Periodicity of the cycle will not be established during the early stages, and the presumptive diagnosis may be totally unrelated to a possible malaria infection.
If the fever does develop a synchronous cycle, it is usually a cycle of somewhat less than 48 h. An untreated primary attack of P. True relapses from the liver do not occur, and after a year, recrudescences are rare. Severe or fatal complications of P. What are some of the problems associated with the differentiation between Plasmodium spp. The ring forms of all four species of Plasmodium can mimic the ring forms of Babesia spp.
Multiple rings per cell are more typical of P. Babesia rings are often numerous, of smaller sizes, and tend to be very pleomorphic, while those of P. Differentiation between Plasmodium spp. Often, the parasitemia in a Babesia infection may be heavier than that seen with P. It is also important to remember that some of the Babesia spp.
What is the significance of finding only ring forms on two sets of blood films drawn 6 h apart? Remember that all of the life cycle stages rings, developing trophozoites, early schizonts, late schizonts, mature schizonts, and gametocytes can be seen on the blood films in infections with P. Due to unique characteristics of the life cycle, only rings and gametocytes and occasional mature schizonts are seen in the peripheral blood with a P.
Therefore, if you see two sets of blood films collected 6 h apart that contain ring forms only, there is an excellent chance the patient is infected with P. What should be considered if the patient has been diagnosed with a P. Although relatively uncommon, infections with the fifth human malaria, P.
Why is it important to identify malaria organisms to the species level? It is also important because of potential drug resistance [chloroquine, P. How should a positive malaria blood film be reported? How should results be reported if Plasmodium spp. It is important to convey to the physician that P. The report should read: How often do mixed infections occur and how should they be reported?
When rings are present, along with other developing stages P. Another report example might be: Plasmodium vivax rings, developing schizonts, and gametocytes; possible mixed infection: Why is it important to report the Plasmodium spp. Often patients who have been diagnosed with Plasmodium spp. Chloroquine will not eliminate any gametocytes present, and there are mosquitoes within the United States that can transmit malaria if they take a blood meal from an individual with gametocytes in the blood.
How do parasitemia and malaria severity correlate? The following percentages are helpful in interpretation of malaria severity:. How should blood films be examined for proficiency testing PT? Since you have no idea what organisms might be present, always review the blood films using the 10X objective entire slide. This examination is likely to reveal any microfilariae that are present; however, small parasites like Plasmodium and Babesia may be missed.
Before reporting the smear as negative, examine at least oil immersion fields using the X oil immersion objective. Do Proficiency Testing PT blood films match those seen from actual patients?
Blood films for PT are actual patient specimens, however, they may have a higher parasitemia than is seen in many patients reporting to the ER, clinic, etc. Often, smears contain a higher parasitemia than is commonly seen. So, the PT smears represent a mix, some of which are fairly typical and some of which have a high number of organisms present.
As in the case of a traveler, the thin blood film may appear to be negative, while the thick film will be positive! In a clinical setting, these blood film results are not that unusual, particularly in an immunologically naïve patient traveler with no prior exposure to Plasmodium spp.
What blood parasites might be seen in PT specimens? The following parasites may be seen in PT specimens: What stains are recommended for staining microfilariae? Also, the sheath of Brugia malayi will stain pink with Giemsa stain. What other methods can be used for the identification of blood parasites? Concentrations trypanosomes, microfilariae, leishmaniae Buffy coat thick and thin blood films all blood parasites, including Plasmodium spp.
The Binax rapid test is as sensitive as a good microscopist; however, often these skilled individuals may not be available on off-hour shifts. However, it is important to recognize the pros and cons of the rapid test when compared to microscopic examination of blood films, particularly in travelers in whom the parasitemia may be considerably less than 0.