been initiated. In cases where the patient is hospitalized, monitoring should be performed at 24, 48, and 72 h after initiating therapy. Generally, the parasitemia drops very quickly within the first 2 h; however, in cases of drug resistance, the level may not decrease but actually may increase over time.
Malarial infections should be reported as the percentage of infected RBCs per 100 RBCs counted (0.5%, 1.0%, etc.) (3). Considering the low parasitemia frequently seen in patients within the United States, several hundred RBCs may have to be counted to arrive at an accurate count and determination of the percentage. The thin blood film must be used for this approach.
Another approach is to count the number of parasites per 100 WBCs on the smear. Either the thick or thin film can be used for this purpose. This figure can be converted to the number of parasites per microliter of blood; divide the number of parasites per 100 WBCs by 100, and multiply that figure by the number of WBCs per microliter of blood. Depending on the parasitemia, 200 or more WBCs may have to be counted, so the denominator may vary (it may be 200 or even more). In this case, blood for both the peripheral smears and cell counts must be collected at the same time.
It is critical that the same reporting method be used consistently for every subsequent set of blood films so that the parasitemia can be tracked for decrease or possible increase, indicating resistance. Also, remember that drug resistance may not become evident for a few days. The parasitemia may initially appear to decline but may then begin to increase after several days. Therefore, it is very important that patient parasitemia be monitored, particularly if an infection with P. falciparum has been diagnosed. Drug resistance has also been reported in P. vivax cases; mixed infections are also much more common than suspected.
Although therapy for Babesia infections is usually quite effective, calculation of parasitemia for positive Babesia infections is highly recommended. Babesiosis usually results in self-cure or cure after administration of azithromycin plus atovaquone or clindamycin plus quinine. Although certain individuals who are severely immunocompromised may not respond to these drug regimens, there has been no reported evidence that treatment failure is due to drug-resistant strains of Babesia microti. Preliminary studies indicate that reduction of pathogen loads and WBC inactivation within blood components using riboflavin and ultraviolet light have been used as an alternative for gamma-irradiation for B. microti, Babesia divergens, T. cruzi, HIV, and bacteria (15).
Diagnosis of Malaria: Review of Alternatives to Conventional Microscopy
It is well known that malaria causes significant morbidity and mortality worldwide, including in countries where imported cases are seen. In many developing countries where malaria is highly endemic, diagnostic testing is often inadequate or unavailable due to a lack of trained personnel or funds or both. Although microscopic examination of stained thick and thin blood films remains the standard of practice, this approach is time-consuming, is based on the need for a great deal of expertise in microscopic morphology, and requires the purchase and maintenance of expensive equipment. Rapid diagnostic tests (RDTs) offer great potential to improve the diagnosis of malaria, particularly in remote areas (World Health Organization. 2012. Malaria rapid diagnostic test performance. Summary results of WHO product testing of malaria RDTs: Round 1–4 [2008–2012]; http://www.who.int/malaria/publications/rdtmalaria_ [accessed 6/20/2013]). This provides very comprehensive lists of product testing to date. There are a number of new approaches to the diagnosis of malaria, including the use of fluorescent stains (QBC), dipstick antigen detection of histidine-rich protein 2 (HRP2), parasite lactate dehydrogenase (pLDH), and Flow anti-pLDH Plasmodium monoclonal antibodies), PCR, and automated blood cell analyzers. Parasitemia and its clinical correlates are given in Table 7.7, while some of the testing options for malaria can be found in Table 7.8. Testing options for some of the other blood parasites can be seen in Table 7.9.
Histidine-rich protein 2 of P. falciparum (PfHRP2) is a water-soluble protein that is produced by the asexual stages and gametocytes of P. falciparum, expressed on the RBC membrane surface, and shown to remain in the blood for at least 28 days after the initiation of antimalarial therapy. Many RDTs are based on the detection of PfHRP2, but reports from field tests have questioned their sensitivity and reliability. However, the variability in the results of PfHRP2-based RDTs may be related to the variability in the target antigen (16). This hypothesis was tested by examining the genetic diversity of PfHRP2, which includes numerous amino acid repeats, in 75 P. falciparum lines and isolates originating from 19 countries and testing a subset of parasites by use of two PfHRP2-based RDTs. There is extensive diversity in PfHRP2 sequences, both within and between countries. Logistic regression analysis indicated that two types of repeats were predictive of RDT detection sensitivity (87.5% accuracy), with predictions suggesting that only 84% of P. falciparum parasites in the Asia-Pacific region are likely to be detected at densities of ≤250 parasites/µl. Data also indicate that PfHRP3 may play a role in the performance of PfHRP2-based RDTs. These findings provide an alternative explanation for the variable sensitivity in field tests of malaria RDTs that is not due to the quality of the RDTs (16).
The persistence of parasite HRP2 in the circulation after parasite clearance has been considered a drawback for RDTs targeting HRP2 and a major cause of false-positive results. In one study when PCR was used as the gold standard rather than microscopy, the high rate of RDT false-positive parasitemia results in comparison with microscopy was shown to predominantly represent cases that had a parasite density below the threshold for detection by microscopy (17). Despite the generally low disease-endemic prevalence of malaria in the area, there was a high prevalence of chronic infections with low, fluctuating parasite densities that were better detected by RDT. In areas known to have low-density parasitemias, RDTs targeting HRP2 may increase the diagnostic sensitivity in comparison with microscopy. While microscopy remains the standard for comparison of the diagnostic accuracy for malaria, the limitations of microscopy, and the possibility that RDTs may have superior accuracy in some circumstances, should be taken into account when interpreting the results of diagnostic trials.
To determine the accuracy of RDTs for ruling out malaria in nonimmune travelers returning from areas where malaria is endemic, 21 studies and 5,747 individuals were surveyed from the published literature (18). Diagnostic accuracy studies of nonimmune individuals with suspected malaria were included if they compared rapid tests with expert microscopic examination or PCR tests. The authors concluded that RDTs for malaria may be a useful diagnostic adjunct to microscopy in centers without major expertise in tropical medicine. Initial decisions on treatment initiation and choice of antimalarial drugs can be based on travel history and posttest probabilities after rapid testing. However, expert microscopy is still required for species identification and confirmation.
Diagnostic accuracy of RDTs for self-diagnosis was variable, and requires improvements before widespread acceptance (19). RDTs have limitations that include the difficulty in detecting mixed infections, all species of Plasmodium, and infections at low concentrations
