Seasonal pattern of Echinococcus re-infection in owned dogs in Tibetan communities of Sichuan, China and its implications for control

Owned dogs are considered to be most important zoonotic transmission source for both
human cystic echinococcosis (CE) and alveolar echinococcosis (AE) on the Qinghai-Tibet
Plateau 10], 11]. A previous study in Shiqu County showed that the faeces of owned dogs were mainly
distributed around the houses of dog owners (proximity 0–200 m) 24]. Faeces from owned dogs that were PCR positive for E. multilocularis DNA were also shown to be spatially clustered 25]. Control of echinococcosis in dogs is therefore an integral component of public health
programme aiming to control human echinococcosis in the region.

A strategy for sustainable control of canine echinococcosis should ideally be based
on an understanding of dog re-infection patterns, especially in areas co-endemic for
CE and AE 26]. A single oral administration of praziquantel (5.0 mg/kg bw) to dogs was found to
be 99.9 % effective against both E. granulosus and E. multilocularis22], 27], 28]. Current Echinococcus spp. copro-antigen ELISA tests are genus specific up to 95 % and thus provide a useful
tool to measure prevalence 29]. Moss et al. 30] used copro-ELISA and copro-PCR to determine re-infection of owned dogs in Shiqu County
after a single dose of praziquantel in May 2006. The study found an average copro-ELISA
prevalence of 10 % two months after this treatment (July 2006); 3 % five months after
(October 2009); and 11 % one year after (May 2007). Despite not using repeated dosing
as in the current study, these results are suggestive of a lower infection pressure
in summer and autumn and higher infection pressure in winter and early spring. The
current research was designed to assess overall Echinococcus spp. re-infection patterns of dogs in Tibetan communities in two counties (Shiqu
and Seda) in Sichuan Province over a 12 month period (April 2009 to April 2010) with
particular attention paid to potential seasonal variation. As such, all dogs were
treated with praziquantel at each gsampling round in order to ensure that any infections
detected at the subsequent round from dosed dogs occurred during the period between
treatment and sampling. The current study also found some evidence of a lower re-infection
pressure in summer and autumn, and a higher re-infection pressure in spring. In addition,
it revealed a more specific higher re-infection pressure season of early winter (from
October to December). The current study did not find an influence on the seasonality
of re-infection by different deworming/sampling time spans for both studies. Early
winter is the main slaughtering season in the region, where the prevalence of livestock
CE has been estimated to be between 7.3 and 76 % for yaks 31]–35]; 14 and 82 % for sheep; and 3.5 and 48 % for goats 36]. The current study therefore suggests that intensive praziquantel treatment of dogs
(preferably monthly dosing) would be advantageous in early winter, spring and early
summer.

Possible death of intermediate hosts of E. granulosus due to extreme cold and forage shortage in winter and early spring was assumed to
be an important risk factor for the prevalence of CE on the plateau 9], 17], 31]. These hosts include wild ungulates and livestock (yaks, sheep and goats). Wild ungulates
have been found to have a prevalence of CE, with estimates of 6.42 % (21/327) amongst
blue sheep (Pseudois nayaur) and 6.57 % (13/198) for Tibetan gazelle (Procapra picticaudata) in the Qinghai part of the Plateau 37]. Therefore, it was considered that a higher opportunity for exposure/re-infection
of E. granulosus in dogs could occur in winter and early spring. However, the current study did not
support the assumption of a higher exposure in the period between December 2009 and
April 2010, which is the winter and early spring in the region. No report of extreme
cold and forage shortage in the period might explain why the assumption was not supported.

Several small mammal species, such as the Qinghai vole (Microtus fuscus) and the Plateau pika (Ochotona curzoniae), are known to act as intermediate hosts for E. multilocularis, with the plateau pika considered to be of particular importance 38], 39]. The plateau pika is known to be diurnal, with most activity observed between 06:30
and 20:00 between January and April (based upon field investigations in January, April,
August, October and November) 19], 20]. Owned tethered dogs were usually released at around 20:00 and leashed again at around
08:00 the following morning – providing these dogs with opportunities to prey on small
mammals with nocturnal and/or crepuscular activity 24], 30]. The current study found that the highest re-infection prevalence occurred in July
2009 (for study A), following dosing in April, which suggests that the highest exposure
to Echinococcus spp. occurred between these two months. This may be associated with greater opportunities
for access to small mammal hosts during spring and early summer.

A preference for keeping male dogs in Tibetan communities has been observed in other
surveys 30], 40], and the same pattern was found in the current study. Despite female dogs being less
commonly kept than male dogs (amongst owned dogs), a significantly higher Echinococcus spp. copro-prevalence was observed amongst female dogs than male dogs at the first
sampling (April 2009). The reason behind this higher prevalence in female dogs is
unknown. It is also not clear why the prevalence in Seda county were higher than those
in Shiqu county during the first two rounds of sampling points (April and July 2009)
(Table 3). These differences were not apparent in subsequent visits.

The Echinococcus re-infection prevalence was not found to be associated with the previous infection
status at any sampling point. The prevalence (1.56 % for both study populations) at
the sampling point in April 2010 were significantly lower than those in April 2009
(13.36 % for study A and 12.5 % for study B) (Fig. 2). The two results might imply the re-infection was largely determined by the existing
reservoir of the parasite in hosts and the availability of the infectious viscera
of intermediate hosts. Thus, it could be assumed that the owned dogs’ role as definite
host to influence the reservoir was reduced drastically during the deworming. Therefore,
it demonstrates the effect of repeated praziquantel dosing on reducing the prevalence
of infection amongst owned dogs.

Stray dogs were not included in the current study. Although the stray dog population
density was not found to be associated with the prevalence of human echinococcosis
in a review of literature published between January 2000 and July 2011 13], the role of stray dogs in the life cycle is not fully understood in the area. Further
work investigating levels of infection amongst stray dogs would therefore be useful.
Also, the number of dogs sampled decreased in each of the five sampling periods for
Study A. This is partly associated with the Tibetan nomadic life style, where people
move with their livestock (yaks and sheep) and dogs to higher pastures between May
and October. Future research to measure the re-infection of owned dogs in summer pastures
would be useful.

Regular deworming of all owned dogs is very hard to apply and sustain in the vast,
remote, high altitude and difficult terrain of eastern Qinghai-Tibet Plateau. A New
Zealand backed pilot intervention project in Garze County for cystic echinococcosis
which included livestock vaccination, was not very successful in large part due to
the logistics of dog deworming in the isolated area 41]. There is therefore a need for a simpler and more sustainable deworming strategy.
It has been argued that effective dosing of owned dogs 2–4 times per year could have
a major impact on both zoonotic risk and transmission potential for both E. multilocularis and E. granulosus in Tibetan communities 30]. One of the criteria defined by WHO for effective control of human CE as a public
health problem, is to reduce the canine echinococcosis prevalence to 0.01 % 42]. The current study demonstrates that the canine prevalence could not be reduced to
this level within one year by with four doses of praziquantel. This is likely due
to the reservoir of infection in intermediate hosts, and suggests that higher dosing
frequencies may be required to achieve low prevalence. The current study also suggests
that ‘targeted’ anthelmintic dosing of dogs during the spring and early winter could
be beneficial.