title: Misclassification of Plasmodium infections by conventional microscopy and the impact of remedial training on the proficiency of laboratory technicians in species identification.
abstract: BACKGROUND
  Malaria diagnosis is largely dependent on the demonstration of parasites in stained blood films by conventional microscopy. Accurate identification of the infecting Plasmodium species relies on detailed examination of parasite morphological characteristics, such as size, shape, pigment granules, besides the size and shape of the parasitized red blood cells and presence of cell inclusions. This work explores misclassifications of four Plasmodium species by conventional microscopy relative to the proficiency of microscopists and morphological characteristics of the parasites on Giemsa-stained blood films.
  
  
  METHODS
  Ten-day malaria microscopy remedial courses on parasite detection, species identification and parasite counting were conducted for public health and research laboratory personnel. Proficiency in species identification was assessed at the start (pre) and the end (post) of each course using known blood films of Plasmodium falciparum, Plasmodium malariae, Plasmodium ovale and Plasmodium vivax infections with densities ranging from 1,000 to 30,000 parasites/μL. Outcomes were categorized as false negative, positive without speciation, P. falciparum, P. malariae, P. ovale, P. vivax and mixed infections.
  
  
  RESULTS
  Reported findings are based on 1,878 P. falciparum, 483 P. malariae, 581 P. ovale and 438 P. vivax cumulative results collated from 2008 to 2010 remedial courses. Pre-training false negative and positive misclassifications without speciation were significantly lower on P. falciparum infections compared to non-falciparum infections (p < 0.0001). Post-training misclassifications decreased significantly compared to pre- training misclassifications which in turn led to significant improvements in the identification of the four species. However, P. falciparum infections were highly misclassified as mixed infections, P. ovale misclassified as P. vivax and P. vivax similarly misclassified as P. ovale (p < 0.05).
  
  
  CONCLUSIONS
  These findings suggest that the misclassification of malaria species could be a common occurrence especially where non-falciparum infections are involved due to lack of requisite skills in microscopic diagnosis and variations in morphological characteristics within and between Plasmodium species. Remedial training might improve reliability of conventional light microscopy with respect to differentiation of Plasmodium infections.
authors.affiliation: Kenya Medical Research Institute/United States Army Medical Research Unit, Kenya, Malaria Diagnostics Centre, Box 54 - 40100, Kisumu, Kenya. peter.obare@usamru-k.org
authors.forename: Peter
authors.initials: P
authors.lastname: Obare
authors.forename: Bernhards
authors.initials: B
authors.lastname: Ogutu
authors.forename: Mohammed
authors.initials: M
authors.lastname: Adams
authors.forename: James Sande
authors.initials: JS
authors.lastname: Odera
authors.forename: Ken
authors.initials: K
authors.lastname: Lilley
authors.forename: David
authors.initials: D
authors.lastname: Dosoo
authors.forename: Christine
authors.initials: C
authors.lastname: Adhiambo
authors.forename: Seth
authors.initials: S
authors.lastname: Owusu-Agyei
authors.forename: Fred
authors.initials: F
authors.lastname: Binka
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Wanja
authors.forename: Jacob
authors.initials: J
authors.lastname: Johnson
mesh_terms: D003951:Diagnostic Errors; D004502:Education, Medical, Continuing; D006801:Humans; D059232:Laboratory Personnel; D008288:Malaria / Q000175:diagnosis* / Q000469:parasitology; D008853:Microscopy / Q000379:methods*; D010274:Parasitology / Q000379:methods*; D010961:Plasmodium / Q000145:classification* / Q000166:cytology*; D011361:Professional Competence; D013194:Staining and Labeling / Q000379:methods
references.citation: Clin Infect Dis. 2011 Jun;52(11):1356-62
references.pmid: 21596677
references.citation: Trends Parasitol. 2007 Jun;23(6):278-83
references.pmid: 17459775
references.citation: Am J Trop Med Hyg. 2007 Dec;77(6 Suppl):119-27
references.pmid: 18165483
references.citation: PLoS Pathog. 2011 Apr;7(4):e1002015
references.pmid: 21490952
references.citation: Malar J. 2013 Jan 08;12:8
references.pmid: 23294844
references.citation: Rev Peru Med Exp Salud Publica. 2010 Oct-Dec;27(4):540-7
references.pmid: 21308193
references.citation: PLoS Med. 2010 Jun 15;7(6):e1000290
references.pmid: 20563310
references.citation: Malar J. 2011 Feb 11;10:38
references.pmid: 21314950
references.citation: Malar J. 2008 Sep 11;7:174
references.pmid: 18783630
references.citation: Am J Trop Med Hyg. 2008 Mar;78(3):442-8
references.pmid: 18337341
references.citation: Acta Trop. 2004 Mar;90(1):61-4
references.pmid: 14739024
references.citation: Lancet Infect Dis. 2009 Sep;9(9):555-66
references.pmid: 19695492
references.citation: J Infect Dis. 2010 May 15;201(10):1544-50
references.pmid: 20380562
references.citation: Malar J. 2009 Apr 21;8:73
references.pmid: 19383118
references.citation: Am J Trop Med Hyg. 2000 Feb;62(2):225-31
references.pmid: 10813477
references.citation: Clin Microbiol Rev. 2007 Oct;20(4):579-92
references.pmid: 17934075