**2. Laboratory test**

#### **2.1. Characteristics of diagnostic tests**

A diagnostic test intends to identify whether or not a patient has a disease that cannot be recognized by signs and symptoms. Ideally, such test must meet the following requirements:


Different studies can be carried out in the dengue research. For example, clinical diagnosis diseases, epidemiological studies, clinical assays, viral load tests, vaccines and anti-viral assessments, etc. Thus, it is necessary to know the testing capacity to detect the presence or absence of the disease *(validation of diagnostic tests)*, so it is very useful to know the following diagnostic test indicators (**Table 1**).


**Sensitivity**: It is the probability that a sick person delivers a positive result in the diagnostic test (a/(a + c)).

**Specificity**: It is the probability that a person who is not sick delivers a negative results in the diagnostic test (d/(b + d)). **Positive predictive value**: It is the probability that a person is sick and gets a positive result in the diagnostic test (a/(a + b)).

**Negative predictive value**: It is the probability that a healthy person gets a negative result in the diagnostic test (d/(c + d)). **Efficiency**: It is the probability that a person is properly diagnosed using the diagnostic test ((a + d)/(a + b + c + d)).

**Positive likelihood ratio (**LR+ **)**: It is obtained when dividing sensitivity by the portion of false positives (1 − specificity), and it indicates the probability of being sick if the result is positive (Sensitivity/(1 − specificity)).

**Negative likelihood ratio (**LR−**)**: It is obtained when dividing false negatives (1 − sensitivity) by specificity, and it indicates the probability of negative results obtained from a sick person ((1 − Sensitivity)/Specificity).

**Table 1.** A double-entry table to obtain indicators of diagnostic tests from analyzing a sample showing all possible results.

The sensitivity and specificity have distinctive features of diagnostic tests, they are not compromised by the prevalence of the disease, and they are inversely proportional. If a study in which the majority of people are suffering from this disease is carried out, a high sensitivity test is needed in order to identify the highest number of true positive and the lowest number of false negative. However, it may increase the number of false positive. If you want to obtain a good disease diagnosis, a high specificity test must be used to detect the highest number of true negative. Here, the false positive will also be low. Moreover, positive and negative predictive values of the diagnostic tests are affected by the prevalence of the disease in the study population. The likelihood ratio (LR) that is independent of prevalence is used when the laboratory tests do not present dichotomous results but cut-off value. This is another way of assessing the accuracy. According to the results, the test can be classified into adequate (LR+ ≥ 10 to LR<sup>−</sup> ≤ 0.1), moderate (LR+ ≥ 5 < 10 to LR<sup>−</sup> > 0.1 ≤ 0.2), scarce (LR+ ≥ 2 < 5 to LR<sup>−</sup> > 0.2 ≤ 0.5) and insignificant (LR+ ≥ 1 < 2 to LR<sup>−</sup> > 0.5 < 1). Not only one but also many tests can be used to diagnose dengue in the epidemiological studies. It can be done sequentially or in parallel. For example, when performing a test with two sequential tests, all positive people need to be assessed with a second test upfront, and this will cause the reduction of net sensitivity and a net specificity enhancement obtained from both tests. It will be considered as positive if their tests are positive in all tests. Likewise, the negative ones will have negative results in the confirmatory test. On the other hand, if two simultaneous tests are used, a net sensitivity is gained, while a net specificity is reduced. This is different when tests are done independently. Negative is considered people whose negative results were in all tests and positive the ones whose positive results were in at least one of the tests [3, 4].

#### **2.2. Biological samples for dengue studies**

and surveillance programs [2]. Laboratory tests are based on the detection of the virus, part of its genome or structure, or specific result from an infected person or animal as immune response. In this chapter, the most used laboratory tests in an arbovirus study focused on diagnosis and research of dengue virus (DENV) will be assessed, some comparisons will be carried out with other laboratory tests, its features, advantages and disadvantages, and cau-

A diagnostic test intends to identify whether or not a patient has a disease that cannot be recognized by signs and symptoms. Ideally, such test must meet the following requirements:

Different studies can be carried out in the dengue research. For example, clinical diagnosis diseases, epidemiological studies, clinical assays, viral load tests, vaccines and anti-viral assessments, etc. Thus, it is necessary to know the testing capacity to detect the presence or absence of the disease *(validation of diagnostic tests)*, so it is very useful to know the following

**Specificity**: It is the probability that a person who is not sick delivers a negative results in the diagnostic test (d/(b + d)). **Positive predictive value**: It is the probability that a person is sick and gets a positive result in the diagnostic test

**Negative predictive value**: It is the probability that a healthy person gets a negative result in the diagnostic test (d/(c + d)). **Efficiency**: It is the probability that a person is properly diagnosed using the diagnostic test ((a + d)/(a + b + c + d)).

**Negative likelihood ratio (**LR−**)**: It is obtained when dividing false negatives (1 − sensitivity) by specificity, and it

**Table 1.** A double-entry table to obtain indicators of diagnostic tests from analyzing a sample showing all possible results.

**Positive a** (True positive) **b** (False positive) **a + b Negative c** (False Negative) **d** (True negative) **c + d Total a + c b + d a + b + c + d Sensitivity**: It is the probability that a sick person delivers a positive result in the diagnostic test (a/(a + c)).

and it indicates the probability of being sick if the result is positive (Sensitivity/(1 − specificity)).

indicates the probability of negative results obtained from a sick person ((1 − Sensitivity)/Specificity).

**Positive Negative Total**

**)**: It is obtained when dividing sensitivity by the portion of false positives (1 − specificity),

tions need to be considered during the process.

58 Dengue Fever - a Resilient Threat in the Face of Innovation

• To achieve high levels of sensitivity and specificity.

**2.1. Characteristics of diagnostic tests**

• Accurate and precise results.

diagnostic test indicators (**Table 1**).

**Result of the laboratory test Disease**

• To deliver rapid results.

• To be cost effective.

(a/(a + b)).

**Positive likelihood ratio (**LR+

**2. Laboratory test**

The type of sample taken in the right moment, storage and transport to the laboratory to be processed, and the appropriate documentation plays a key role to obtain results because if there is a change of sample quality, this can reduce antibody titers, viruses or genetic material resulting in lower titers or concentrations from the real ones in quantitative tests or false negative results in quantitative or qualitative tests.

The most used samples to diagnose and to search about dengue are whole blood, serum, plasma, and human organs like spleen, liver, and heads of mosquitoes, pools of mosquitoes, brains of mice, serum samples saturated with filter paper, etc. The serum samples that will be processed for virus isolation and/or polymerase chain reaction tests and fluorescent focus assay to quantify the virus in serum are collected in tubes without anticoagulants. They must be taken within 1–5 days after the onset of symptoms, to detect the antigen (NS1 protein), within 1–6 days to detect IgM antibodies for enzyme-linked immunosorbent assay (ELISA) or a rapid test after day 5 of starting the symptoms, within 1–5 to detect IgG using matched serums to assess seroconversion for ELISA, neutralization test or hemagglutination inhibition used in acute serums, and convalescent serums after day 15. In the case of liver, spleen, kidney or nodes samples, a sample is immediately taken after the person's death or the animal in experiment. The samples must be taken to the laboratory as soon as possible and preferably dealt with in dry ice or liquid nitrogen [5, 6].
