INDIA /BRAZIL / SOUTH AFRICA TRI-LATERALCOLLABORATIVE PROGRAM ON SCIENCE AND TECHNOLOGYII Forum IBSA on Malaria, Teresina – PI, BR, March, 7th - 8th, 2006

Hernando A del Portillo, PhD Departamento de ParasitologiaInstituto de Ciências Biomédicas Universidade de São Paulo

http://clinmaldb.usp.br/hadelportillo

Diagnosis, Cell and Molecular Biology

DIAGNOSIS

 

Giemsa-staining Malaria Diagnosis Brazil 1960-2005

External Evaluation of Giemsa-staining diagnosis in reference laboratories from the State of Pará

Dra. Marinete Póvoa

• State of Pará more than 100.000 cases/2005• 11 participating laboratories • Five different reference slides: P. falciparum, P.

vivax, mixed infection, P. vivax + T. cruzi, negative control

• 73% of the labs diagnosed correctly all slides, remaining 27% miss-diagnosed 1 slide.

• None of the labs identified T. cruzi.• 5 labs have adequate training; 4 labs require re-

training and 2 labs require capacitance.

Multi-Center Assesstment of Immuno-chromatographic Methods

Coordinated: Prof. Cor Jésus Fernandes Fontes

Total No of Patients: 539 Resuts given by Manaus, Porto Velho, Macapá e Cuiabá.

Giemsa Staining

F V N Total

F 87 0 2 89

OPTIMAL V* 0 104 0 104 N 4 9 333 346

Total 91 113 335 539

* = Positive results of other humana malaria parasite species, here in considered positive for P. vivax

Sensitivity P.falciparum = 95,6% Sensitivity P.vivax = 92,0 Specificity P.falciparum = 99,5% Specificity P.vivax = 100,0% Overall sensitivity for malaria diagnosis = 93,6% Overall specificity for malaria diagnosis = 99,4%

PCR

NESTED-PCR

NESTED-PCR

Evaluation of negativation and detection of three human malaria parasite species: Marinete Póvoa

Blood bank surveys Ricardo Machado

MF Ferreira-Da-Cruz

Mixed Infections Graça Alecrim

Asymptomatics LH Pereira da Silva

Nested-PCR and Malaria Diagnosis at FMT/AM

Nested-PCR detected 14.5% of mixed infections as opossed to 1,8% detected by Giemsa staining

Asymptomatic patients

ELISA

Determination of the cutoff value for IgG-ELISA-P. vivax MSP1-19, using the Receiver Operating Characteristic (ROC) Curve.

Sandra do Lago Moraes-Avila & Antonio Walter Ferreira

Nogueira P, Alves F et al

Giemsa-staining Malaria Diagnosis Brazil 1960-2005

MOLECULAR AND CELL BIOLOGY

 

Polymorphims of Molecular Markers - Parasites

P. vivax Duffy Binding Protein (de Brito, Moreira, Carvalho)

P. falciparum reticulocyte binding-like homologues (PfRh1-4) (Mariano Zalis)

P126 protein (Ferreira-da-Cruz, Banic, Carvalho, Ribeiro)

Glycophorin B (GBP), HFE (Ricardo Machado)

MSP1, MSP2 (Marcelo Urbano)

Baum et al PLoS Path 2005

PfRH1 Polymorphism amino acid # as in HB3

Isolates 2830

2831

2832

2849

2850

(co

de)

2851

2852

2853

HB3 V HNx9 (H N) x9 (Q N) X1 Q K DDd2,3D7,IPN,NS,PSS1, GOS,LANA,JSL V (H N) x4 (Q N) x1 (10D) Q K D7G8,BHZ,21Q, ALR,04Q V (H N) x6 (Q N) x2 (4D) Q K DRPN V (H N) x4 (Q N) x3 (6D) Q K DGVM V (H N) x5 (Q N) x2 (6D*) Q K DFFS,35Q V (H N) x3 (Q N) x2 (10D*) Q K D

PfRH2a Polymorphism

Isolates 2548

2699 to

2713

2734

2735

2736

2737

2738

2739

2740

2741

(co

de)

2803 to

2852

7G8,35Q D no Del K K E A L K K Q (pepA) 50 aa DelHB3,RPN,21Q,GVM,BHZ,04Q,ALR,FFS, JSL D 15 aa^ Del K K E A L K K Q (pepA) 50 aa DelDd2,NS,GOS,IPN, PSS1,LANA D 15 aa Del Q K E E E L K R Q (pepB) 50 aa Del3D7 A 15 aa Del K K E E E L R K K (pepC) no Del

amino acid # as in 7G8

Zalis, M (unpublished)

Microssatelites

Plasmodium vivax

Cor Jesús Fernandes Fontes, Alvaro Couto, Luzia H Carvalho Cristiana FA Brito

2006

Polymorphisms - Human Molecular Markers and Associations with Malaria

Infection

CR-1, MBL, TLR, NOS2, TNF-a, IFN-b, IL-4, IL-10CD36, ICAM-1, Cytochrome P450, GST (Sandra do Lago Moraes)

CSP

MSP119

Mosquitoes and cDNA libraries

W. P. Tadei cDNA library of A. darlingi

oligoCy3

Pf genomic DNA

50% DMSO

salmon sperm DNA

polydA DNA

50% DMSO

Block1=3000, 3000, 100, 22, 194, 22, 194"

columnsrows

oligoCy3

Pf genomic DNA

50% DMSO

salmon sperm DNA

polydA DNA

50% DMSO

Block1=3000, 3000, 100, 22, 194, 22, 194"

columnsrows