Description: Wallach’s Interpretation of Diagnostic Tests, now in its Ninth Edition, has been completely revised and updated by a new author team from the Department of Hospital Laboratories, UMass Memorial Medical Center faculty, who are carrying on the tradition of Jacques Wallach’s teachings. This text serves as a practical guide to the use of laboratory tests which aids physicians in using tests more effectively and efficiently by offering test outcomes, possible meanings, differential diagnosis, and summaries of tests available.

The book has been reorganized into 2 sections. The first section is devoted to an alphabetical listing of laboratory tests while stressing the integration of the clinical laboratory in the clinical decision making process. Test sensitivity, specificity, and positive and negative infectious disease probabilities are included whenever appropriate. Microbiology tests are listed in a separate chapter. The second section is devoted to disease states. Where appropriate, a patient’s chief complaint and/or physical findings are initially presented with subsequent discussions focused on discrete disease states as they relate to a patient’s chief complaint. Current molecular diagnostic testing, cytogenetics, common pitfalls, test limitations, and identification of appropriate tests for specific clinical presentations are also addressed.

Ninth Edition highlights include:
Detailed listing and description of routine and esoteric tests listed alphabetically, with information on when to order and how to interpret the test results based on evidence-based laboratory medicine.
Information on how to work up patients with specific symptoms and the appropriate lab tests to order
Up-to-date test procedures including molecular diagnostic tests
Detailed microbiology chapter of infectious diseases

Spanish health authorities said on Thursday they were investigating a possible outbreak of Crimean-Congo hemorrhagic fever (CCHF) which has killed one man and infected a nurse, in the first non-imported case reported in Western Europe.

The 62-year-old man died on Aug. 25 after contracting the CCHF disease during a walk in the Castilla-Leon region, probably from a tick bite he reported - which is one of the main ways it is transmitted - authorities said in a statement.

He also infected the nurse who treated him at a hospital in Madrid and she is now in a stable condition in quarantine at an isolation unit, they said. Authorities are monitoring about 200 other people who had come into contact with the man and nurse.

According to the World Health Organization, CCHF's mortality rate is about 30 percent and it is endemic to Africa, the Balkans and Ukraine, the Middle East and Central Asia.

Read more: First Local Case of Tick-Borne Disease Kills Man in Spain
Since some species exhibit periodicity (i.e. circulation of microfilariae in increased numbers at certain times of the day), blood should be collected at the correct time to improve the chances of detection. For Wuchereria bancrofti and Brugia malayi showing nocturnal periodicity, blood should be collected at night between 10 p.m. to 4 a.m. Microfilariae are present in greater numbers in capillary blood than in venous blood; therefore, skin puncture is preferred. Usually microfilariae are scanty in peripheral blood so that concentration techniques may be necessary for their demonstration. Following microscopic methods can be used for detection of microfilariae in peripheral blood:
• Thick blood smear
• Concentration techniques: mem-brane filtration, microhematocrit centrifugation, lysed venous blood technique, lysed capillary blood technique.
Thick blood smear
A thick blood smear is spread from 20 μl of capillary blood on a glass slide, air-dried, and stained with a Romanowsky stain. If microfilariae are not detected in thick smears prepared from capillary blood collected at the appropriate time, and if clinical suspicion is strong, concentration techniques are employed. This is because circulating microfilariae are often scanty and sensitivity of microfilarial detection increases when volume of blood sampled is increased.
Brugia Malayi
Concentration techniques
Membrane filtration: This is a sensitive method but is expensive for routine use in endemic areas. Anticoagulated venous blood (10 ml) is passed through a polycarbonate membrane filter of 3 μm or 5 μm pore size. Following this 10 ml of methylene blue saline solution is passed through the filter for staining the microfilariae. Microfilariae are trapped and retained on the filter, which is placed on a glass slide and examined under the microscope.
Microhematocrit tube or capillary tube method: Two heparinised capillary tubes are filled with blood from skin punctures (or two plain capillary tubes are filled with anticoagulated venous blood). After sealing the dry ends with a suitable sealant, tubes are centrifuged in a microhematocrit centrifuge for about 5 minutes. The capillary tubes are placed on a glass slide and fixed with adhesive tape. Plasma just above the buffy coat layer is examined for motile microfilariae under the microscope.
Lysed venous blood method: 10 ml of venous blood is lysed by saponin-saline solution. The hemolysate is centrifuged, supernatant is discarded, and the sediment is placed on glass slide. After adding a drop of methylene blue solution, a coverslip is placed, and the preparation is examined under the microscope for microfilariae.
Lysed capillary blood method: 0.1 ml of blood obtained by skin puncture is added to 1 ml of saponin-saline solution to cause lysis of red cells. After centrifugation, supernatant is discarded and sediment is placed on a glass slide. A drop of methylene blue solution is added and a coverslip is placed over it. The entire preparation is examined under the microscope for motile microfilariae.
Morphology of Microfilariae on Romanowsky-stained Blood Smears Wuchereria bancrofti: Microfilariae measure about 300 μ in length and 8 μ in breadth. They have a hyaline sheath, which stains pink. There are distinct nuclei in the central axis of the body. Nuclei are not present in the tip of the tail. Cephalic space (present at the anterior end) is as long as it is broad. Tip of the tail is bent backwards and body curves are few.
Brugia malayi: These measure about 230 μ × 6 μ in size. Sheath stains dark pink in color. The nuclei are crowded in the body, are blurred in outline, and the tip of the tail shows two distinct nuclei. Cephalic space is twice as long as it is broad. Instead of smooth curves to the body, there are kinks.
Malaria parasites can be detected by identification of specific nucleic acid sequences in their DNA. Methods based on polymerase chain reaction (PCR) have been developed for identification of DNA of malaria parasite. Species diagnosis is also possible. PCR-based methods can detect very low levels of parasites in blood (< 5 parasites/μl of blood) with very high sensitivity and specificity.
Molecular methods can be useful in the diagnosis of malaria, in following response to treatment, in epidemiological surveys, and for screening of blood donors. They can also be used as a standard to judge other methods of malaria diagnosis. However, these methods cannot be routinely applied because of the high cost, need for special equipments and materials, and lengthy procedure (24 hours). Presently they can be used as a research tool in malaria control programs, and to carry out quality control checks on microscopic diagnosis.
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Life history of malaria parasite consists of two cycles of development: asexual cycle or schizogony that occurs in humans and sexual cycle or sporogony that occurs in mosquitoes.

(1) Asexual cycle (human cycle, schizogony): This occurs in the liver cells and red blood cells of infected humans, and therefore humans are the intermediate hosts of the malaria parasite (Schizogony refers to the process of reproduction in protozoa in which there is production of daughter cells by fission). The human cycle begins when infected female Anopheles mosquito bites a person and sporozoites are injected into the circulation. There are four stages of human cycle.
(a) Pre-erythrocytic schizogony (Hepatic schizogony):
Inoculated sporozoites rapidly leave the circulation to enter the liver cells where they develop into hepatic (pre-erythrocytic) schizonts (Schizonts are cells undergoing schizogony). One sporozoite produces one tissue form. Hepatic schizonts rupture to release numerous merozoites in circulation (Merozoites are daughter cells produced after schizogony). Up to 40,000 merozoites are produced in the hepatic schizont.
In P. falciparum infection, all of the hepatic schizonts mature and rupture simultaneously; dormant forms do not persist in hepatocytes. In contrast, some of the sporozoites of P. vivax and P. ovale remain dormant after entering liver cells and develop into schizonts after some delay. Such persistent forms are called as hypnozoites; they develop into schizonts at a later date and are a cause of relapse.
(b) Erythrocytic schizogony:
Merozoites released from rupture of hepatic schizonts enter the red blood cells via specific surface receptors. These merozoites become trophozoites that utilize red cell contents for their metabolism. A brown-black granular pigment (malaria pigment or hemozoin) is produced due to breakdown of hemoglobin by malaria parasites. The fully formed trophozoite develops into a schizont by multiple nuclear and cytoplasmic divisions. Mature schizonts rupture to release merozoites, red cell contents, malarial toxins, and malarial pigment. (This pigment is taken up by monocytes in peripheral blood and by macrophages of reticulo-endothelial system. In severe cases, organs which are rich in macrophages like spleen, liver, lymph nodes, and bone marrow become slate-gray or black in color due to hemozoin pigment). Rupture of red cell schizonts corresponds with clinical attack of malaria. Released merozoites infect new red cells and enter another erythrocytic schizogony cycle. This leads to rapid amplification of plasmodia in the red cells of the human host. In P. falciparum, P. vivax, and P. ovale infections, cycle of schizogony lasts for 48 hours, while in P. malarie infection it lasts for 72 hours. Merozoites of P. vivax and P. ovale preferentially invade young red cells or reticulocytes while those of P. falciparum infect red cells of all ages. Senescent red cells are preferred by P. malariae.
P. vivax, P. ovale, and P. malariae complete the erythrocyte schizogony in general circulation. Schizonts of P. falciparum induce membrane changes in red cells, which causes them to adhere to the capillary endothelial cells (cytoadherence). Therefore, in P. falciparum infection, erythrocyte schizogony is completed in capillaries of internal organs and usually only ring forms are seen in circulation.
(c) Gametogony:
After several cycles of erythrocytic schizogony, some merozoites, instead of developing into trophozoites and schizonts, transform into male and female gametocytes. These sexual forms are infective to mosquito and the person harboring them is called as a “carrier”. Gametocytes are not pathogenic for humans.
(d) Exoerythrocytic schizogony:
In P. vivax and P. ovale infections, some of the sporozoites in liver cells persist and remain dormant. These dormant forms in liver cells are called as hypnozoites. They become active and develop into schizonts a few days, months, or even years later. These schizonts rupture, release merozoites, and cause relapse. Exoerythrocytic schizogony is absent in P. falciparum infection and therefore relapse does not occur. Hence, P. vivax and P. ovale are called as relapsing plasmodia while P. falciparum and P. malariae are known as non-relapsing plasmodia.

(2) Sexual cycle (mosquito cycle, sporogony): The sexual cycle begins when a female Anopheles mosquito ingests mature male and female gametocytes during a blood meal. First, 4-8 microgametes are produced from one male gametocyte (microgametocyte) in the stomach of the mosquito; this is called as exflagellation. The female gametocyte (macrogametocyte) undergoes maturation to produce one macrogamete. By chemotaxis, microgametes are attracted toward the macrogamete; one of the microgametes fertilizes the macrogamete to produce a zygote. The zygote becomes motile and is called as ookinete. Ookinete penetrates the lining of the stomach and comes to lie on the outer surface of the stomach where it develops into an oocyst. On further growth and maturation, multiple sporozoites are formed within the oocyst. After complete maturation, oocyst ruptures to release sporozoites into the body cavity of the mosquito. Most of the sporozoites migrate to the salivary glands. Infection is transmitted to the humans by the bite of the mosquito through saliva when it takes a blood meal.


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