during october 12 , 2014march 28 , 2015 , samples collected from 1,058 persons in liberia with suspected ebov infection were submitted to the cdc  nih elwa laboratory .
the samples used for this research were collected for public health surveillance and not human subjects research , so institutional review board review and approval were not required .
early during the study period ( october  november ) , most of the patients from whom samples were collected received a diagnosis of ebov infection ( figure 1 , panel a ) ; toward the end , however , few cases of ebov infection were diagnosed in liberia . the number of patients who received a diagnosis of plasmodium spp .
parasitemia remained stable over time ; thus , despite the lack of positive ebov test results in the final months of the study period , overall , 40%60% of patients each week received a diagnosis of ebov infection , plasmodium spp .
parasitemia , or both ( figure 1 , panel a ) . of 1,058 samples tested , 259 ( 24.5% ) were positive for ebov alone , 243 ( 23% ) were positive for plasmodium spp .
alone , and 47 ( 4.4% ) were positive for both ( figure 1 , panel b ) .
of 311 plasmodium - positive samples that were further analyzed , 296 ( 95% ) were positive for p. falciparum ( figure 1 , panel c ) , confirming that p. falciparum was the main plasmodium species causing parasitemia in our cohort .
rna in patient samples submitted to the centers for disease control and prevention  national institutes of health diagnostic laboratory at the eternal love winning africa campus in monrovia , liberia , from october 12 , 2014 ( epidemiologic week 42 ) , through march 28 , 2015 ( week 13 ) .
whole blood samples were inactivated , and rna was extracted by using the qiaamp viral rna mini kit ( qiagen , hilden , germany ) .
rna by real - time quantitative reverse transcription pcr ( qrt - pcr ) ( 13 ) .
a ) number of patients , as determined by qrt - pcr , positive for ebov , plasmodium spp .
, both , or neither ( i.e. , no ebov and no plasmodium spp . ) , by epidemiologic week .
b ) total number of patients receiving a laboratory diagnosis of ebola viremia , plasmodium spp .
qrt - pcr  positive samples that were retested with a qrt - pcr specific for p. falciparum ( 14 ) .
a high ct value corresponds to a low - level parasitemia ; the lower the ct value , the higher the number of plasmodium parasites detected .
all patients in the cohort were triaged as having suspected ebov infection , and thus all had clinical symptoms that might have been caused by ebov , plasmodium spp . , or a different pathogen . of note
, significantly fewer plasmodium parasites were detected in patients with ebov infection than in patients who were not infected with ebov ( average plasmodium ct 24.7 vs. 20.37 ; p<0.01 by unpaired student t - test ) , likely because clinical symptoms in these patients were caused by the ebov infection rather than malaria . some of the patients in our cohort with plasmodium spp .
parasitemia might have been asymptomatic carriers , especially those with a low - level parasitemia .
parasitemia over time by month of sample submission , for samples submitted to the centers for disease control and prevention  national institutes of health diagnostic laboratory at the eternal love winning africa campus in monrovia , liberia , from october 12 , 2014 , through march 28 , 2015 .
cycle threshold ( ct ) values were detected by using real - time quantitative reverse transcription pcr .
triangles represent parasite loads in parasitemic patients co - infected with ebola virus ; circles represent patients with plasmodium spp .
similar to other malaria - endemic regions , a large proportion of the population is infected with plasmodium parasites without developing clinical disease . to compensate for this and the higher sensitivity of the pcr assay compared with light microscopy
, we used a cutoff of ct 
 < 30 rather than ct 
 < 40 under the assumption that a 10-ct difference would compensate for the 1,000-fold higher sensitivity of pcr over microscopy .
this principle could be carried further to assume that a ct 
 < 25 would be in the range detectable by the rapid diagnostic test . of note
one could argue that because all patients with febrile illness who were seen at many of the etus were given antimalarial treatment when they first sought medical care , providing laboratory testing for plasmodium spp .
presumptive artemisinin - based combination treatment is recommended for all patients seen at etus , followed by prompt malaria diagnostic testing so that appropriate measures can be taken if oral treatment can not be sustained because of clinical symptoms ( 9,10 ) . moreover ,
mathematical models predict a large increase in malaria in the epidemic region because of lapses in malaria control ( 7 ) .
parasitemia testing of all febrile patients seen at healthcare facilities could be used to determine whether this increase is indeed occurring so that countermeasures can be scaled up accordingly .
differential diagnostic testing should be expanded to identify the cause of disease in the patients in our cohort whose samples tested negative for ebov and plasmodium spp .
, as was attempted in 1 laboratory in sierra leone ( 15 ) . however , this kind of testing was difficult for several reasons .
first , during the height of the evd epidemic in liberia , most laboratories were already working close to capacity while testing for ebov alone . testing for additional pathogens would have required the allocation of additional resources , including equipment and personnel .
second , it might not be possible to perform diagnostics using a whole blood sample for all the pathogens of differential diagnostic importance that can cause signs and symptoms similar to those of evd ( e.g. , typhoid , bacterial sepsis , shigellosis , cholera , leptospirosis , dengue fever , rickettsioses , relapsing fever , meningitis , viral hepatitis , influenza , lassa fever ) .
third , each of these pathogens likely is the cause of disease in only a small subset of febrile patients .
because broad - spectrum antimicrobial drugs and antimalarial drugs are routinely administered to all patients seen at an etu , consideration for additional testing should focus on endemic pathogens that would not respond to these treatments . by using pcr - based detection for plasmodium spp .
parasitemia , the need for additional handling of clinical specimens possibly infected with ebov was eliminated .
thus , the addition of pcr - based diagnostic tests to detect plasmodium spp . does not pose an additional safety risk to laboratory staff .
also , it is less time - consuming to add additional pcr reactions to a pcr run than to separately perform microscopy or an rdt on each sample , and these additions would add only 15 minutes to the overall time needed from sample submission to reporting of results .
moreover , if a multiplex approach is used , the additions would not require extra time or reagents .
parasitemia can be implemented easily and safely in laboratories performing ebov diagnostics to assist with case - patient management during evd outbreaks in malaria - endemic areas .
 malaria is a major public health concern in the countries affected by the ebola virus disease epidemic in west africa . 
 we determined the feasibility of using molecular malaria diagnostics during an ebola virus disease outbreak and report the incidence of plasmodium spp . 
 parasitemia in persons with suspected ebola virus infection . 
The Study
Conclusions