Diagnosis of placental malaria in poorly fixed and processed placental tissue.

BACKGROUND: Placental histopathology has been considered the gold standard for diagnosis of malaria during pregnancy. However, in under-resourced areas placental tissue is often improperly fixed and processed; the resulting formalin pigment is difficult to distinguish from malaria pigment. This study examines two alternative diagnostic methods: polymerase chain reaction (PCR) and a novel immunohistochemistry (IHC)-based method using an antibody against histidine-rich protein 2 (HRP2). METHODS: Placental histopathology from 151 pregnant women in Kinshasa was assessed by two blinded microscopists and compared with peripheral blood PCR and IHC for HRP2. The Cohen's kappa coefficients were calculated to assess the test agreement. The sensitivity and specificity of individual tests were calculated using PCR or IHC as the reference standard as well as latent class analysis (LCA). RESULTS: PCR and IHC correlated fairly well. The correlation between the two blinded microscopists was poor, as there was widespread formalin pigment. Using LCA, all of the tests had high specificities. The most sensitive test was IHC (67.7 %), with PCR as second-best (56.1 %). CONCLUSIONS: PCR and/or IHC are suitable diagnostics when the presence of formalin pigment substantially compromises placental histopathology.

Direct evidence of extensive diversity of HIV-1 in Kinshasa by 1960.

Human immunodeficiency virus type 1 (HIV-1) sequences that pre-date the recognition of AIDS are critical to defining the time of origin and the timescale of virus evolution. A viral sequence from 1959 (ZR59) is the oldest known HIV-1 infection. Other historically documented sequences, important calibration points to convert evolutionary distance into time, are lacking, however; ZR59 is the only one sampled before 1976. Here we report the amplification and characterization of viral sequences from a Bouin's-fixed paraffin-embedded lymph node biopsy specimen obtained in 1960 from an adult female in Léopoldville, Belgian Congo (now Kinshasa, Democratic Republic of the Congo (DRC)), and we use them to conduct the first comparative evolutionary genetic study of early pre-AIDS epidemic HIV-1 group M viruses. Phylogenetic analyses position this viral sequence (DRC60) closest to the ancestral node of subtype A (excluding A2). Relaxed molecular clock analyses incorporating DRC60 and ZR59 date the most recent common ancestor of the M group to near the beginning of the twentieth century. The sizeable genetic distance between DRC60 and ZR59 directly demonstrates that diversification of HIV-1 in west-central Africa occurred long before the recognized AIDS pandemic. The recovery of viral gene sequences from decades-old paraffin-embedded tissues opens the door to a detailed palaeovirological investigation of the evolutionary history of HIV-1 that is not accessible by other methods.