Limits...
Real-time PCR: Benefits for Detection of Mild and Asymptomatic Giardia Infections.

Prasertbun R, Sukthana Y, Popruk S - Trop Med Health (2012)

Bottom Line: Children who live in crowded conditions or low socio-economic areas are the risk group for Giardia infection.The detection limit of real-time PCR for G. duodenalis was 0.1 pg/25 µl reaction.It can detect both mild and asymptomatic Giardia infections in children living on the Thai-Myanmar border.

View Article: PubMed Central - PubMed

Affiliation: Department of Protozoology, Faculty of Tropical Medicine, Mahidol University, Bangkok 10400, Thailand.

ABSTRACT
The majority of Giardia infections are transmitted by the fecal-oral route and cause giardiasis. Children who live in crowded conditions or low socio-economic areas are the risk group for Giardia infection. Interestingly, most of them are asymptomatic or only mildly infected and can shed the Giardia cysts in the environment. Thus, the diagnosis of Giardia infection in asymptomatic or mild infection plays an important role in achieving control of Giardia duodenalis transmission. The objective of this study was to examine parasitic infections using microscopy and to develop a real-time PCR method for detection of Giardia infection in the stool samples of children living on the Thai-Myanmar border. Both species-specific primers and fluorescent labeled G. duodenalis probe were designed using small-subunit ribosomal RNA (ssrRNA). The results showed that 10 (7.69%) and 40 (30.77%) of 130 stool samples were positive for G. duodenalis by microscopy and real-time PCR respectively. Only 3 out of 9 liquid stools revealed G. duodenalis positive using microscopy, but all of them were G. duodenalis-positive using real-time PCR. The detection limit of real-time PCR for G. duodenalis was 0.1 pg/25 µl reaction. It can detect both mild and asymptomatic Giardia infections in children living on the Thai-Myanmar border.

No MeSH data available.


Related in: MedlinePlus

Standard curve of 10-fold serial dilution of Giardia duodenalis trophozoite DNA. 1 = 10 ng, 2 = 1 ng, 3 = 0.1 ng, 4 = 0.01 ng, 5 = 1 pg, 6 = 0.1 pg; R = 0.997
© Copyright Policy - open-access
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3475312&req=5

Figure 3: Standard curve of 10-fold serial dilution of Giardia duodenalis trophozoite DNA. 1 = 10 ng, 2 = 1 ng, 3 = 0.1 ng, 4 = 0.01 ng, 5 = 1 pg, 6 = 0.1 pg; R = 0.997

Mentions: In this study, all Giardia positive samples were calculated directly based on DNA weight in nanogram from the standard curve (Fig. 3). Moreover, the number of trophozoites or cysts in Giardia positive stool samples was calculated by the following equation: X ng/0.144 and X ng /0.313, respectively (X: Quantity of G. duodenalis DNA sample obtained from the standard curve; each individual G. duodenalis trophozoite and mature cyst contained ~0.144 pg and 0.313 pg of DNA, respectively) [12]. In fact, it is difficult to differentiate the original source of these DNA of G. duodenalis (trophozoite or cyst stage). Thus, we assumed that the number of G. duodenalis in stools can be calculated from the cyst stage. The results revealed that liquid stools contained higher amounts of G. duodenalis than soft or hard stools (liquid stool, 500–2,000 cysts/25 µl of reaction; soft stool, 75–400 cysts/25 µl of reaction; and hard stool, 6–300 cysts/25 µl of reaction), as shown in Table 2.


Real-time PCR: Benefits for Detection of Mild and Asymptomatic Giardia Infections.

Prasertbun R, Sukthana Y, Popruk S - Trop Med Health (2012)

Standard curve of 10-fold serial dilution of Giardia duodenalis trophozoite DNA. 1 = 10 ng, 2 = 1 ng, 3 = 0.1 ng, 4 = 0.01 ng, 5 = 1 pg, 6 = 0.1 pg; R = 0.997
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3475312&req=5

Figure 3: Standard curve of 10-fold serial dilution of Giardia duodenalis trophozoite DNA. 1 = 10 ng, 2 = 1 ng, 3 = 0.1 ng, 4 = 0.01 ng, 5 = 1 pg, 6 = 0.1 pg; R = 0.997
Mentions: In this study, all Giardia positive samples were calculated directly based on DNA weight in nanogram from the standard curve (Fig. 3). Moreover, the number of trophozoites or cysts in Giardia positive stool samples was calculated by the following equation: X ng/0.144 and X ng /0.313, respectively (X: Quantity of G. duodenalis DNA sample obtained from the standard curve; each individual G. duodenalis trophozoite and mature cyst contained ~0.144 pg and 0.313 pg of DNA, respectively) [12]. In fact, it is difficult to differentiate the original source of these DNA of G. duodenalis (trophozoite or cyst stage). Thus, we assumed that the number of G. duodenalis in stools can be calculated from the cyst stage. The results revealed that liquid stools contained higher amounts of G. duodenalis than soft or hard stools (liquid stool, 500–2,000 cysts/25 µl of reaction; soft stool, 75–400 cysts/25 µl of reaction; and hard stool, 6–300 cysts/25 µl of reaction), as shown in Table 2.

Bottom Line: Children who live in crowded conditions or low socio-economic areas are the risk group for Giardia infection.The detection limit of real-time PCR for G. duodenalis was 0.1 pg/25 µl reaction.It can detect both mild and asymptomatic Giardia infections in children living on the Thai-Myanmar border.

View Article: PubMed Central - PubMed

Affiliation: Department of Protozoology, Faculty of Tropical Medicine, Mahidol University, Bangkok 10400, Thailand.

ABSTRACT
The majority of Giardia infections are transmitted by the fecal-oral route and cause giardiasis. Children who live in crowded conditions or low socio-economic areas are the risk group for Giardia infection. Interestingly, most of them are asymptomatic or only mildly infected and can shed the Giardia cysts in the environment. Thus, the diagnosis of Giardia infection in asymptomatic or mild infection plays an important role in achieving control of Giardia duodenalis transmission. The objective of this study was to examine parasitic infections using microscopy and to develop a real-time PCR method for detection of Giardia infection in the stool samples of children living on the Thai-Myanmar border. Both species-specific primers and fluorescent labeled G. duodenalis probe were designed using small-subunit ribosomal RNA (ssrRNA). The results showed that 10 (7.69%) and 40 (30.77%) of 130 stool samples were positive for G. duodenalis by microscopy and real-time PCR respectively. Only 3 out of 9 liquid stools revealed G. duodenalis positive using microscopy, but all of them were G. duodenalis-positive using real-time PCR. The detection limit of real-time PCR for G. duodenalis was 0.1 pg/25 µl reaction. It can detect both mild and asymptomatic Giardia infections in children living on the Thai-Myanmar border.

No MeSH data available.


Related in: MedlinePlus