Utilidad de la prueba Aspergillus-LFD para el diagnóstico de aspergilosis: primera experiencia en Chile / Utility of Aspergillus-LFD: first experience in Chile

Introduction: Invasive fungal diseases (IFD) by filamentous fungi are a common cause of morbidity and mortality in immunocompromised patients, especially those with myeloid leukemia. In 2011 a protocol for the rapid diagnosis of IFD by filamentous fungi was implemented in Valparaiso Region. Objectives: To describe cases of IFD by filamentous fungi of the Valparaíso Region, since the implementation of rapid diagnosis and to compare results with the period 2004-2009. Materials and Method: Descriptive and prospective study conducted in two public hospitals: Carlos van Buren at Valparaiso and Gustavo Fricke at Viña del Mar. We selected patients with a diagnosis of filamentous fungal diseases considering the EORTC/MSG criteria. Demographics, underlying diseases, risk factors for EFI, galactomannan (GM) results in blood and bronchoalveolar lavage, cultures and biopsies, treatment and overall lethality rates at 30 days were registered. Results: Eighteen patients were detected, 6 with proven and 12 probable IFD. Nine were diagnosed by GM, 8 by culture and two with both methods. In cases which the agent (9/18) was isolated from Rhizopus oryzae was the most frequent. When comparing overall lethality with the period 2004-2009, there was a reduction of 47.8%, which was statistically significant. Conclusions: Compared to data previously published in the region, demographic and comorbidities of patients with IFD caused by filamentous fungi are similar, however the currently rapid diagnosis protocol has improved survival of patients and lethality experienced overall decrease.

Introducción: la enfermedad fúngica invasora (EFI) por hongos filamentosos es una causa frecuente de morbilidad y mortalidad en pacientes inmunocomprometidos, en especial en aquellos con leucemia mieloide. En el 2011 se implementó en la Región de Valparaíso un protocolo de diagnóstico rápido de la EFI por hongos filamentosos. Objetivos: describir los casos de EFI por hongos filamentosos de la Región de Valparaíso, desde la implementación del diagnóstico rápido y compararlos con el período 2004-2009. Materiales y Método: Estudio descriptivo y prospectivo realizado en los hospitales públicos Carlos van Buren de Valparaíso y Gustavo Fricke de Viña del Mar. Se seleccionaron aquellos pacientes con diagnóstico de EFI por hongos filamentosos considerando los criterios EORTC/MSG. Se obtuvieron datos demográficos, enfermedad de base, factores de riesgo para EFI, resultados de galactomanano (GM), cultivos y biopsias, tratamiento y letalidad global a 30 días. Resultados: Se identificaron 18 pacientes, seis con EFI probadas y 12 probables. Nueve fueron diagnosticados con galactomanano, ocho con cultivos y uno con los dos métodos. En los casos en que se aisló el agente (9/18), Rhizopus oryzae fue el más frecuente. Al comparar la letalidad global con la del período 2004-2009, hubo una reducción de 47,8%, la cual fue estadísticamente significativa. Conclusiones: En relación a lo publicado anteriormente en la región, se conservan las características demográficas y de co-morbilidad de los pacientes con EFI por hongos filamentosos; sin embargo, la introducción del nuevo protocolo de diagnóstico rápido se asoció a una disminución en la letalidad global.

A robust HIV-1 viral load detection assay optimized for Indian sub type C specific strains and resource limiting setting

BACKGROUND: Human Immunodeficiency Virus Type 1 (HIV-1) viral load testing at regular intervals is an integral component of disease management in Acquired Immunodeficiency Syndrome (AIDS) patients. The need in countries like India is therefore an assay that is not only economical but efficient and highly specific for HIV-1 sub type C virus. This study reports a SYBR Green-based HIV-1 real time PCR assay for viral load testing and is designed for enhanced specificity towards HIV-1 sub type C viruses prevalent in India. RESULTS: Linear regression of the observed and reference concentration of standards used in this study generated a correlation coefficient of 0.998 (p<0.001). Lower limit of detection of the test protocol was 50 copies/ml of plasma. The assay demonstrated 100% specificity when tested with negative control sera. The Spearman coefficient of the reported assay with an US-FDA approved, Taqman probe-based commercial kit was found to be 0.997. No significant difference in viral load was detected when the SYBR Green based assay was used to test infected plasma stored at -20°C and room temperature for 7 days respectively (Wilcoxon signed rank test, p=0.105). In a comparative study on 90 pretested HIV-1 positive samples with viral loads ranging from 5,000 - 25,000 HIV-1 RNA copies/ml and between two commercial assays it was found that the later failed to amplify in 13.33% and 10% samples respectively while in 7.77% and 4.44% samples the copy number values were reduced by >0.5 log value, a figure that is considered clinically significant by physicians. CONCLUSION: The HIV-1 viral load assay reported in this study was found to be robust, reliable, economical and effective in resource limited settings such as those existing in India. PCR probes specially designed from HIV-1 Subtype C-specific nucleotide sequences originating from India imparted specificity towards such isolates and demonstrated superior results when compared to two similar commercial assays widely used in India.

Cost-effective analysis of different algorithms for the diagnosis of hepatitis C virus infection

We compared the cost-benefit of two algorithms, recently proposed by the Centers for Disease Control and Prevention, USA, with the conventional one, the most appropriate for the diagnosis of hepatitis C virus (HCV) infection in the Brazilian population. Serum samples were obtained from 517 ELISA-positive or -inconclusive blood donors who had returned to Fundação Pró-Sangue/Hemocentro de São Paulo to confirm previous results. Algorithm A was based on signal-to-cut-off (s/co) ratio of ELISA anti-HCV samples that show s/co ratio ≥95 percent concordance with immunoblot (IB) positivity. For algorithm B, reflex nucleic acid amplification testing by PCR was required for ELISA-positive or -inconclusive samples and IB for PCR-negative samples. For algorithm C, all positive or inconclusive ELISA samples were submitted to IB. We observed a similar rate of positive results with the three algorithms: 287, 287, and 285 for A, B, and C, respectively, and 283 were concordant with one another. Indeterminate results from algorithms A and C were elucidated by PCR (expanded algorithm) which detected two more positive samples. The estimated cost of algorithms A and B was US$21,299.39 and US$32,397.40, respectively, which were 43.5 and 14.0 percent more economic than C (US$37,673.79). The cost can vary according to the technique used. We conclude that both algorithms A and B are suitable for diagnosing HCV infection in the Brazilian population. Furthermore, algorithm A is the more practical and economical one since it requires supplemental tests for only 54 percent of the samples. Algorithm B provides early information about the presence of viremia.

Determining cost-effectiveness and cost component of three malaria diagnostic models being used in remote non-microscope areas.

This cross-sectional experimental study developed a methodology to analyze the cost-effectiveness of three malaria diagnostic models: microscopy; on-site OptiMAL; and on-site Immunochromatographic Test (on-site ICT), used in remote non-microscope areas in Thailand, from both a public provider and patient perspective. The study covered six areas in two highly malaria-endemic areas of provinces located along the Thai-Myanmar border. The study was conducted between April and October 2000, by purposively recruiting 436 malaria suspected cases attending mobile malaria clinics. Each patient was randomly selected to receive service via the three diagnostic models; their accuracy was 95.17%, 94.48% and 89.04%, respectively. In addition, their true positive rates for all malaria species were 76.19%, 82.61% and 73.83%; for falciparum malaria 85.71%, 80.95% and 80.00%, and for vivax malaria 57.14%, 100% and 50%, respectively, with the parasitemia ranging from 80 to 58,240 microl of blood. Consequently, their costs were determined by dividing into provider and consumer costs, which were consequently classified into internal and external costs. The internal costs were the costs of the public providers, whereas the external costs were those incurred by the patients. The aggregate costs of these three models were 58,500.35, 36,685.91, and 40,714.01 Baht, respectively, or 339.53, 234.39, and 243.93, in terms of unit costs per actual case. In the case of microscopy, if all suspected malaria cases incurred forgone opportunity costs of waiting for treatment, the aggregate cost and unit cost per actual case were up to 188,110.89 and 944.03 Baht, respectively. Accordingly, the cost-effectiveness for all malaria species, using their true positive rates as the effectiveness indicator, was 446.75, 282.40, and 343.56 respectively, whereas for falciparum malaria it was 394.80, 289.37 and 304.91, and for vivax malaria 595.67, 234.39 and 487.86, respectively. This study revealed that the on-site OptiMAL was the most cost-effective. It could be used to supplement or even replace microscopy for this criteria in general. This study would be of benefit to malaria control program policy makers to consider using RDT technology to supplement microscopy in remote non-microscope areas.

New, simple and cheap alternative to troponin test for diagnosis of acute myocardial infarction.

Acute myocardial infarction (AMI) is often a fatal disorder in humans seen throughout the world. It was earlier diagnosed with some serum enzymes like aspartate transaminase, creatine phosphokinase and its isoenzyme CPK-MB and lactate dehydrogenase which were shown to be increased in AMI. However, in the last few years importance has been given to measuring serum troponins released from the injured myocardium to confirm an AMI. Troponin estimation involves immunological technique, which is expensive with other associated problems like shelf life of reagents, number of samples to be analysed and availability of the kit itself, used for estimation. Under these circumstances the present work involves the measurement of total salt soluble proteins which are proteins associated with troponins also released from myocardium of a patient with AMI. This new test overrules all the disadvantages of the troponin test but seems equally viable and useful for diagnosis of AMI.

Ex post and ex ante willingness to pay (WTP) for the ICT Malaria Pf/Pv test kit in Myanmar.

Willingness to pay (WTP) for the ICT Malaria Pf/Pv test kit was assessed by the contingent valuation method using a bidding game approach in two villages in Myanmar. Kankone (KK) village has a rural health center (RHC) and Yae-Aye-Sann (YAS) is serviced by community health worker (CHW). The objectives were to assess WTP for the ICT Malaria Pf/Pv test kit and to determine factors affecting the WTP. In both villages WTP was assessed in two different conditions, ex post and ex ante. The ex post WTP was assessed at an RHC in the KK village and at the residence of a CHW in the YAS village on patients immediately following diagnosis of malaria. The ex ante WTP was assessed by household interviews in both villages on people with a prior history of malaria. Ordinary least squares (OLS) multiple regression analysis was used to analyze factors affecting WTP. The WTP was higher in ex post conditions than ex ante in both villages. WTP was significantly positively associated with the average monthly income of the respondents and severity of illness in both ex post and ex ante conditions (p < 0.001). Distance between the residence of the respondents and the health center was significantly positively associated (p < 0.05) in the ex ante condition in a household survey of YAS village. Traveling time to RHC had a negative relationship with WTP (p < 0.05) in the ex post condition in the RHC survey in KK village.

A comparison of three malaria diagnostic tests, under field conditions in North-west Thailand.

A hospital-based trial to compare the clinical diagnosis of malaria; microscopy, and a rapid diagnostic antigen capture detection dipstick (ParaSight-F) was conducted in North-west Thailand. 301 people who presented themselves at the hospital were selected. 204 (68%) were presumptively diagnosed as having malaria by the triage nurses; 64 (21.3%) were P. falciparum parasite positive, and 94 (32%) tested positive for P. falciparum with the ParaSight-F test strips. There was no association between hemoglobin levels (<10g/dl and > or = 10g/dl) and malaria, and although there was a good statistical association between temperature and malaria the specificity, sensitivity and positive predictive values were all low, indicating that temperature alone is a poor indicator of the disease. Based on the microscopy results, we found that a presumptive clinical diagnosis dramatically over-diagnosed malaria, and similarly there were a large number of false positives using the ParaSight-F test. We believe that many of the patients had received some form of malaria treatment prior to presentation at the hospital, and that the high number of false positives are explained by persistent antigenemia and the possibility of there being sequestered parasites following incomplete chemotherapy.

An indigenously developed nitrite kit to aid in the diagnosis of urinary tract infection.

OBJECTIVE: To evaluate the utility of an indigenously developed nitrite kit for the rapid diagnosis of urinary tract infection (UTI) METHODS: 1018 urine specimens were collected from all cases where there was clinical suspicion of UTI. Samples were cultured as per standard microbiological protocol. Presence of nitrites was indicated by the development of purple color on addition of color developing solution and compared with the set of graded positive and negative controls also provided in the Kit. RESULTS: The results of the nitrite kit were compared with the semi-quantitative urine culture as the gold standard. The sensitivity, specificity, positive predictive and negative predictive values were 47%, 87%, 31% and 93%, respectively. CONCLUSION: Nitrite kit as a screening test can decrease the work load in the clinical bacteriology laboratory. More importantly in a field set up that is devoid of culture facilities, it can be used to correctly predict the absence of UTI.

Assessing the economic impact of a rapid on-site malaria diagnostic test.

A set of three models has been developed for assessing the economic impact of existing and new malaria diagnostic technology, specifically microscopy of blood slides and rapid on-site diagnostic tests (RDT). The models allow for phased introduction of the new technology in targeted areas. The derived computer software program facilitates evaluation of costs to the supplier, to the consumer and aggregate costs, with comparison among the three models to give relative costs of progressive transition from blood slides to RDT technology. The models and the related software program can assist planners in the health sector in determining costs of current programs and assessing the potential economic impact of introducing rapid on-site diagnosis. Details of the models and the operational software program are available on request.