Sensitivity of nested-PCR for plasmodium detection in pooled whole blood samples and its usefulness to blood donor screening in endemic areas.

Transfusion-transmitted malaria is a severe disease with high fatality rate. Most Brazilian blood banks in the Amazon region perform malaria screening using microscopic examination (thick smears). Since low parasite concentrations are expected in asymptomatic blood donors a high sensitivity test should be used for donor screening. This study determined the sensitivity of a nested-PCR for plasmodium detection in pooled samples. We performed a one-stage criterion validation study with 21 positive samples pooled with samples from ten negative volunteer until three different concentrations were reached (0.33; 0.25; 0.20 parasites/µL - p/µL). Nested PCR was performed as described by Snounou et al. (1993). Sensitivities (and confidence intervals) were determined by stratum of final parasite concentration on the pooled samples. All samples with parasitemia values of 0.33 and 0.25 p/µL had 100% sensitivity (95%CI=86.3-100). One negative result was obtained from a sample with 0.20 p/µL sensitivity=95.2% (95%CI=76.2-99.9). Compared to parasitemia detectable under ideal conditions of thick smear, this nested-PCR in pooled sample was able to detect 40 times more parasites per microliter. Nested-PCR in pooled samples should be considered as a high sensitive alternative to thick smear for donor screening in blood banks at endemic regions. Local authorities need to assess cost:benefit advantages of this method compared to alternatives.

Barriers with Valve Mechanisms Are Predicted to Protect Crops from Slug Carriers of Rat Lungworm Disease.

Angiostrongyliasis (Rat Lungworm disease) is an emerging parasitic disease caused by the ingestion of gastropods infected with the neurotropic nematode Angiostrongylus cantonensis. The reduction of crop infestation with infected slug carriers may vary widely by protection method. We explored the application of barriers with valve mechanisms, whereby selective directional forces caused a greater number of slugs to exit than enter the protected plot, leading to decreased slug population densities at a steady state. Using field data, we constructed predictive models to estimate slug population densities at a steady state in protected plots with (1) no valve effect, (2) a valve effect, (3) no valve effect with a single breach of the barrier, (4) a valve effect with a single breach of the barrier, (5) a valve effect with a constant breach of the barrier, and (6) a repelling effect. For all scenarios, plots protected using a barrier with a valve effect had consistently lower slug densities at a steady state. Our findings support the use of barriers with valve mechanisms under different conditions, and potentially in combination with other interventions to reduce the contamination of crops by slug carriers of A. cantonensis. Improving barriers extends beyond disease mitigation to economic and cultural impacts on the local farmer and consumer communities.

Diagnostic approach to acute diarrheal illness in a military population on training exercises in Thailand, a region of campylobacter hyperendemicity.

High rates of Campylobacter fluoroquinolone resistance highlight the need to evaluate diagnostic strategies that can be used to assist with clinical management. Diagnostic tests were evaluated with U.S. soldiers presenting with acute diarrhea during deployment in Thailand. The results of bedside and field laboratory diagnostic tests were compared to stool microbiology findings for 182 enrolled patients. Campylobacter jejuni was isolated from 62% of the cases. Clinical and laboratory findings at the time of presentation were evaluated to determine their impact on the posttest probability, defined as the likelihood of a diagnosis of Campylobacter infection. Clinical findings, the results of tests for inflammation (stool occult blood testing [Hemoccult], fecal leukocytes, fecal lactoferrin, plasma C-reactive protein), and the numbers of Campylobacter-specific antibody-secreting cells in peripheral blood failed to increase the posttest probability above 90% in this setting of Campylobacter hyperendemicity when these findings were present. Positive results by a Campylobacter-specific commercial enzyme immunoassay (EIA) and, less so, a research PCR were strong positive predictors. The negative predictive value for ruling out Campylobacter infection, defined as a posttest probability of less than 10%, was similarly observed with these Campylobacter-specific stool-based tests as well the fecal leukocyte test. Compared to the other tests evaluated, the Campylobacter EIA is a sensitive and specific rapid diagnostic test that may assist with diagnostic evaluation, with consideration of the epidemiological setting, logistics, and cost.

Traveler's diarrhea in Thailand: randomized, double-blind trial comparing single-dose and 3-day azithromycin-based regimens with a 3-day levofloxacin regimen.

BACKGROUND: Traveler's diarrhea in Thailand is frequently caused by Campylobacter jejuni. Rates of fluoroquinolone (FQ) resistance in Campylobacter organisms have exceeded 85% in recent years, and reduced fluoroquinolone efficacy has been observed. METHODS: Azithromycin regimens were evaluated in a randomized, double-blind trial of azithromycin, given as a single 1-g dose or a 3-day regimen (500 mg daily), versus a 3-day regimen of levofloxacin (500 mg daily) in military field clinics in Thailand. Outcomes included clinical end points (time to the last unformed stool [TLUS] and cure rates) and microbiological end points (pathogen eradication). RESULTS: A total of 156 patients with acute diarrhea were enrolled in the trial. Campylobacter organisms predominated (in 64% of patients), with levofloxacin resistance noted in 50% of Campylobacter organisms and with no azithromycin resistance noted. The cure rate at 72 h after treatment initiation was highest (96%) with single-dose azithromycin, compared with the cure rates of 85% noted with 3-day azithromycin and 71% noted with levofloxacin (P=.002). Single-dose azithromycin was also associated with the shortest median TLUS (35 h; P=.03, by log-rank test). Levofloxacin's efficacy was inferior to azithromycin's efficacy, except in patients with no pathogen identified during the first 24 h of treatment or in patients with levofloxacin-susceptible Campylobacter isolates, in whom it appeared to be equal to azithromycin. The rate of microbiological eradication was significantly better with azithromycin-based regimens (96%-100%), compared with levofloxacin (38%) (P=.001); however, this finding was poorly correlated with clinical outcome. A higher rate of posttreatment nausea in the 30 min after receipt of the first dose (14% vs. <6%; P=.06) was observed as a mild, self-limited complaint associated with single-dose azithromycin. CONCLUSIONS: Single-dose azithromycin is recommended for empirical therapy of traveler's diarrhea acquired in Thailand and is a reasonable first-line option for empirical management in general.

Failure of national guidelines to diagnose uncomplicated malaria in Bangladesh.

During the mid 1990s, national guidelines were established in accordance with World Health Organization recommendations for the diagnosis of uncomplicated malaria in Bangladesh. Based on simple clinical and epidemiologic criteria these guidelines were designed to be applied outside of tertiary care centers where microscopy was not feasible. We evaluated the positive predictive value (PPV) of these criteria using microscopic slide examinations as the gold standard in 684 subjects diagnosed and treated for malaria, sampling from eight subdistrict centers. The PPV for malaria was 32% with 19% for falciparum and 14% for Plasmodium vivax. Medical officers assigned to the study also gave their own clinical impression of whether cases could have been malaria. With the additional criteria of a medical officers' diagnosis, the PPV increased negligibly to 37% with 23% and 14% for falciparum and vivax, respectively. Since the PPV of diagnosis is low and cannot be improved on clinical grounds alone, we recommend the incorporation of laboratory diagnosis. This is especially important as we detect resistance to the first-line therapy chloroquine and require more expensive, potentially more toxic, regimens.

Proguanil plus sulfamethoxazole in the treatment of uncomplicated Plasmodium falciparum malaria.

In a malaria endemic area of Brazil where P. falciparum is highly resistant to chloroquine and Fansidar, we conducted an in vivo study to evaluate the therapeutic response of proguanil plus sulfametoxazole against Plasmodium falciparum malaria. Twenty-five adult subjects with uncomplicated P. falciparum malaria received supervised drug administration and were followed for 28 days in an inpatient hospital or in a malaria free-transmission area. The therapeutic regimen was proguanil 100 mg BID plus sulfamethoxazole 1,000 mg BID for 7 days. Of those who took all medications (n=21), 17 (81%) were cured. Recrudescent parasitemia during follow-up occurred in four (19%) patients on days 14, 19, 20 and 21 after beginning of treatment. The remaining four (16%) subjects did not complete their therapeutic regimen because the incidence of side effects. Considering the shortage of falciparum malaria therapeutic options and the urgent need for new regimens to deal with the spread of drug resistant P. falciparum, one might consider the study results as a lead to study analogous compounds, hopefully with fewer adverse reactions.

Effects of Upcountry Maui water additives on health.

Since 2001 Upcountry Maui residents have been concerned that water additives may be linked to health problems in their community. A study using phone surveys was conducted to assess this issue. Most people suffered skin rashes, while others experienced eye irritations or respiratory problems. The surveys suggested that these symptoms might have been attributable to the water additives.

Antimalarial drug susceptibility testing of Plasmodium falciparum in Brazil using a radioisotope method

From March 1996 to August 1997, a study was carried out in a malaria endemic area of the Brazilian Amazon region. In vivo sensitivity evaluation to antimalarial drugs was performed in 129 patients. Blood samples (0.5 ml) were drawn from each patient and cryopreserved to proceed to in vitro studies. In vitro sensitivity evaluation performed using a radioisotope method was carried out with the cryopreserved samples from September to December 1997. Thirty-one samples were tested for chloroquine, mefloquine, halofantrine, quinine, arteether and atovaquone. Resistance was evidenced in 96.6 percent (29/30) of the samples tested for chloroquine, 3.3 percent (1/30) for quinine, none (0/30) for mefloquine and none for halofantrine (0/30). Overall low sensitivity was evidenced in 10 percent of the samples tested for quinine, 22.5 percent tested for halofantrine and in 20 percent tested for mefloquine. Means of IC 50 values were 132.2 (SD: 46.5) ng/ml for chloroquine, 130.6 (SD: 49.6) ng/ml for quinine, 3.4 (SD: 1.3) ng/ml for mefloquine, 0.7 (SD: 0.3) ng/ml for halofantrine, 1 (SD: 0.6) ng/ml for arteether and 0.4 (SD: 0.2) ng/ml for atovaquone. Means of chloroquine IC 50 of the tested samples were comparable to that of the chloroquine-resistant strain W2 (137.57 ng/ml) and nearly nine times higher than that of the chloroquine-sensitive strain D6 (15.09 ng/ml). Means of quinine IC 50 of the tested samples were 1.7 times higher than that of the low sensitivity strain W2 (74.84 ng/ml) and nearly five times higher than that of the quinine-sensitive strain D6 (27.53 ng/ml). These results disclose in vitro high resistance levels to chloroquine, low sensitivity to quinine and evidence of decreasing sensitivity to mefloquine and halofantrine in the area under evaluation