mercado de trabajo
2.6.3. Condiciones de trabajo de los inmigrantes en la construcción
Non-infected 30 0 2 30 28 - -
1-20days 30 4 2 26 28 13.3 6.7
21-34days 20 10 8 10 12 50 40
35-53days 16 16 15 0 1 100 94
Total 96 30 27 66 69 45.5 42.4
Table 5.2(b): Statistical result for comparison of coproantigen ELISA and IGCA rapid test
ELISA & ICA
Time courses with ELISA
Time courses with ICA
Chi-square 0.099 56.592 54.696
P 0.753 0.000 0.000
5.3.3 Copro PCR results for experimental infected dogs
A total of 158 DNA samples were extracted from feces from above E. granulosus
experimental infected dogs. The positive rate was not so good since the fecal sample had stored for near 8 yrs ago. But the interesting result was the clear positive zones occurred in sample No 93 and No. 155 which were fecal samples from two 20-day infected dogs (Fig. 5.6).
Fig. 5.6: No. 93# and 155# showed all positives within double check with Copro PCR (Abbasi’s methods) for experimental infected dogs.
Lanes: fist and last lane were 100-bp molecular DNA ladder, second and third lanes from right were negative and positive control, the other lanes were different DNA samples from dog faeces.
(a) No. 93# (20 days post infection) had one positive lane in 133bp and No. 155# (20 days post infection) had another strong bands.
(b) No. 93# had two obvious positive bands and NO. 155# had a weak positive in its second band.
5.4 Discussion
The dog is a key host in the life cycle of Echinococcus, not just for E. granulosus,
but also for E. multilocularis since it has been confirmed to be infected with E. multilocularis (Craig et al., 1992, 2000, 2003, 2005; Shi, 1995; Vuitton et al., 2003).
Serological tests for dog infection were effective but not so sensitive because serum antibodies may be present from recent infection even if there is no current infection (Craig, 1997). Eggs as the infective source for livestock and human, exist within segments of adult worms and in dog feces. Eggs are however difficult to be differentiated from other taeniid cestode eggs under the microscope. Arecoline purgation is useful to show presence of adult worms after purgation and has been applied in mass screening (Gemmell, 1973; Craig et al., 1995; Budke et al., 2005; Lahmar et al., 2007, 2009). However unsuccessful purgation might cause missed infections and light infections may not be detected (Craig et al., 1995; Schantz et al., 1995; Fraser et al., 2002; Budke et al. 2005). The autopsy of dogs is a gold standard for infection but can not usually be accepted or recommended (Craig et al., 1997; Eckert et al., 1997; WHO/OIE, 2001; Fraser et al, 2002).
Marker N cont ro l P c on tro l 15 8 15 7 15 6 15 5 15 4 15 3 15 2 15 1 10 0 99 98 97 96 95 94 93 92 91 90 89 N cont Marker ro l P c on tro l 15 8 15 7 15 6 15 5 15 4 15 3 15 2 15 1 10 0 99 98 97 96 95 94 93 92 91 90 89 (a) (b)
Coproantigen sandwich ELISA has been widely accepted as a laboratory detection method for canine echinococcosis (Allan et al., 1992; Craig et al, 1995; Deplazes et al, 1992, 1997; WHO/OIE, 2001; Fraser et al, 2002). One of the most common used antigens for raising antibodies was whole worm extract of E. granulosus (EgWWE) and the sensitivity and specificity was generally over 85%
(Allan et al, 1992; Deplazes et al., 1992, 1999). The extracts of E. granulosus
include somatic protoscoleces, excretoary / secretory (E/S) extract of E. granulosus protoscoleces, and the FPLC fraction from EgWWE were all used for
find a more specific antigen to be used as a boost for coproantigen ELISA (Allan et al., 1992; Craig et al., 1995, 1996; Deplazes 1992, 1999; Sakai et al., 1995; Frazer et al., 2002; Elayoubi et al., 2004). Monoclonal antibody for EgWWE was also another attempt to gain more specific result (Kohno et al., 1991, 1995; Sakai et al., 1995; Malgor et al., 1997; Zhang et al., 2003). Copro-PCR was developed for differential diagnosis of E. granulosus and E multilocularis by amphyzation of
species specific DNA (Dinkel et al, 1998; Frazer et al., 2002; Abbasi, 2003; Boufana et al, 2008, 2012).
This Chapter reports the first initial development and assessment of a rapid IGCA for coproantigen detection for effective screening of dog infections. The sensitivity of IGCA was not as good (94%) compared with coproantigen ELISA (100%) in 16 experimental 35-53 dpi dogs. This might be due to several possibilities. Polyclonal antibody was used as the capture and detection antibody, but a good monoclonal antibody was difficult to obtain against a purified antigen. Then the detection system was very important, IGCA is a rapid test, based on eye-reading so its sensitivity and specificity were generally lower than for coproantigen ELISA. Thus it might be best used as an initial screening tool in the field.
The E. granulosus experimental time-course infection of dogs was subjected to
faecal screening. ELISA and IGCA showed that coproantigen level appeared to increase from day 16 after infection and were clearly detected after day 24 dpi by both ELISA and IGCA. This might be thought to correlate when the first segment of worm formed in that period (i.e. day 14-17) and two segmented worm (day 20-28) (WHO/OIE, 2001). And the DNA of E. granulosus was amplified from dog
faeces from 20 days after infection. Copro DNA has been detected in experimentally infected dogs in other studies pre-patency (Lahmar et al., 2010). DNA might be associated with cell turnover from the worm surface and/or lost /
degraded worms or proglottids from 20-28 dpi.
The copro ELISA was also applied to screen faeces of dog sampled in endemic communities in Xinjiang and Qinghai. The initial trial had been used in some community studies and a mean positive rate of 23% (41/178) was obtained. The samples from Hobokersaier and Hejing showed relative higher positives (47.4% and 22.5%) compared to Habahe and Yushu Counties, and also we confirmed the higher human CE prevalence in epidemiological studies in these two counties in Xinjiang (see Chapter 6).
These results suggested that copro tests (coproantigen ELISA, IGCA, copro-PCR) might be sensitive for canine echinococcosis from 20 days after infection. The use of monoclonal antibody for surface antigen of E. granulosus adult worm had been
shown to have effective results in other research work (Zhang et al., 2003) and this could be helpful for both coproantigen ELISA and IGCA. And the IGCA might be potential.
5.5 Summary
A rapid IGCA of coproantigen detection for dog infections was initially developed and assessed. The sensitivity of IGCA was 94% in 16 experimental 35-53 dpi dogs. ELISA and IGCA showed that coproantigen level appeared to increase from day 16 after infection and were clearly detected after day 24 dpi by both ELISA and IGCA. The copro ELISA was also applied to screen faeces of dog sampled in endemic communities in Xinjiang and Qinghai. The initial mean positive rate of 23% (41/178) was obtained. The samples from Hobokersaier and Hejing showed relative higher positives (47.4% and 22.5%).The results suggested that copro tests (coproantigen ELISA, IGCA, copro-PCR) might be sensitive for canine echinococcosis from 20 days after infection.