Past and present of cystic echinococcosis in Bolivia

Viable eggs of the canine intestinal tapeworm Echinococcus granulosus sensu lato (s.l.) infect various intermediate hosts causing cystic echinococcosis (CE). Furthermore, CE represents a serious zoonosis causing a significant global burden of disease. CE is highly endemic in South America, including Argentina, Brazil, Chile, Uruguay, and Peru. For Bolivia, no official data concerning the incidence in humans or the number of livestock and dogs infected are available. However, it is well known that CE occurs in Bolivia. We aim here to fill the gap in the current knowledge of the epidemiological situation of CE in Bolivia, providing a historical overview of documents published within the country, which have never been comprehensively reviewed. The very first documentation of E. granulosus infection in animals dates in 1910, while the first human case was reported in 1913. In total, 876 human CE cases have been reported in the scientific literature, with an apparent increase since the 1970s. In the absence of other epidemiological studies, the highest prevalence in human comes from Tupiza, Potosí Department, where 4.1% (51/1,268) of the population showed signs of CE at mass ultrasound screening in 2011. In the same report, 24% of dog faecal samples were positive for coproantigens of E. granulosus s.l. in ELISA. The highest prevalence in intermediate hosts reported at abattoir reached 37.5% in cattle from Potosí, followed by 26.9% in llamas from Oruro, 2.4% in pigs and 1.4% in sheep from La Paz. Finally, Echinococcus granulosus sensu stricto (s.s.), Echinococcus ortleppi (G5), and Echinococcus intermedius (G7) have been identified in Bolivia. Data reviewed here confirm that E. granulosus s.l. is circulating in Bolivia and that a proper prospective nationwide epidemiological study of CE is urgently needed to define transmission patterns as a basis for the planning and implementation of future control measurements.


Introduction
Cystic echinococcosis (CE), caused by Echinococcus granulosus sensu lato (s.l), is highly endemic in regions of Argentina, Brazil, Chile, Peru, and Uruguay [1]. Substantial research on different aspects of the epidemiology of CE in these countries has been published. a1111111111 a1111111111 a1111111111 a1111111111 a1111111111 Initially diagnosed as lung cancer [39] 1969 Santa Cruz ND ND

ND ND
Lungs Lungs X-ray X-ray ND ND [40] 1971 Oruro Female 20 Lungs X-ray, surgery ND Simultaneous TBC [31] 1973 Santa Cruz Male 5 Lungs X-ray ND 25 cm diameter [41] 1987 US citizen who lived shortly in La Paz  [54]. The majority of the cases visited in these retrospectives studies originated in La Paz, followed by Potosí [57,59]. In the case of the retrospective study by Guzman [55], many patients diagnosed in Tupiza (Potosí) had relatives who previously received surgical treatment for CE. The latter agrees with the high prevalence of humans in Tupiza (4.1%) in 2011 [62]. The other 4 publications mentioned several CE cases without supplying detailed information (not included in Table 2). First, Perez Fontana [63] indicated that Bolivia's representative in a meeting of PAHO reported 64 CE cases; some of these cases were diagnosed in Santa Cruz. Maldonado [31] mentioned in the discussion of a case report that 83 CE cases without concomitant TBC were surgically treated at the Thorax National Institute without supplying a time frame in which these cases were reported. Finally, in 2 reports for PAHO [3,4], Schantz mentioned that 75 surgeries were due to CE out of 1,500 thorax surgeries in 10 years in La Paz. Ultrasound diagnosis, treatment, and patient follow-up. The most accurate and recent data related to human CE cases in Bolivia are found in a government report that showed the

PLOS NEGLECTED TROPICAL DISEASES
First review of cystic echinococcosis in Bolivia results of an ultrasound mass screening performed in Tupiza (Potosí Department). In total, 52 out of 1,268 individuals (4.1%) showed CE's ultrasound signs with active cysts in all patients.
Additional thoracic X-ray showed CE in the lungs of 5 of the same patients with abdominal CE [62]. Patients were treated with albendazole 10 to 15 mg/kg/day for 4 months, and 92% completed the treatment. The follow-up showed that 60% of cysts were classified as transitional at the end of the first month after treatment. By the end of the therapy, 90% of the cysts were classified as inactive [64]. Immunological testing of the Bolivian population. We found only a few comments related to immunodiagnostic tests in humans in the literature. Schantz [3,4] briefly mentioned high reactivity in an intradermic test (with an antigen from E. granulosus) in employees of an abattoir in the Bolivian Altiplano. Commercial [65] and in-house ELISA [66] tests have been used to assess sensitivity and specificity using sera from confirmed CE cases from La Paz and Cochabamba and patients with other parasitic diseases. However, there was no attempt to perform a prevalence study in the general population. As part of the ultrasound study in Tupiza (mentioned above), ELISA and immunochromatography tests (Vircell) were used to detect antibodies against E. granulosus in the sera of 52 patients diagnosed with abdominal CE. In total, 96.7% of the ultrasound cases were positive in both ELISA and immunochromatography. Finally, a study carried out at the Parasitology, Tropical Medicine, and Environment Unit of the Instituto de Investigación en Salud y Desarrollo (IINSAD-UMSA) determined a CE prevalence of 6.3% (44/696) in population from the northern highland at the Department of La Paz using 2 commercial ELISA kits (Nova Tec Inmunodiagnostica and RIDASCREEN) [67].

E. granulosus infection in dogs
Data regarding prevalence in dogs in Bolivia are scarce; all reports of dog infection are summarised in Table 3. Fig 1 shows the Departments of Bolivia for which studies detecting E. granulosus in faeces have been performed. The oldest report of E. granulosus infection in dogs dates from 1973 Schantz [7], where 50% of dogs from La Paz were found infected after arecoline purgation. Methodologies to correctly identify E. granulosus have been used only in 5 publications: Barba [68] did not find positives in the necropsy of 100 dogs from Santa Cruz; Perez [69] did not find infections in 60 dogs purged with arecoline; Villena [70] found adult specimens of E. granulosus in the faeces of 14 out of 85 dogs (16.5%) treated with praziquantel in Tupiza; Subieta [71] determined a prevalence of 12.3% (45/367) in dogs from Tupiza by necropsy; and finally, Ali and colleagues [8] described 4.5% (6/131) of the dogs, from the Altiplano of La Paz, to be infected with E. granulosus s.s. or E. ortleppi based on PCR and sequencing. Other reports showed results based on coproantigen detection in faeces [62,72], a method known to produce cross-reactions with Taenia species, or microscopic detection of taeniid eggs [73][74][75], a technique that does not allow differentiation between Echinococcus and Taenia spp. From the reports using a coproantigen ELISA, Villena determined 24% (63/264) of dog faecal samples as positive with ratios varying between 15% and 62% in different localities of Tupiza (at the border with Argentina) [62]. Subsequently, also using coproantigen ELISA, Casas and colleagues [72] described 3.4% of dogs positive in urban samples from Villazón. In comparison, 30% were positives in the samples taken from a rural area (Lampaya) near the border with Argentina. On the other hand, using microscopic detection of taeniid eggs, Salinas [73] reported 1% (1/100) of dog samples as positive in La Paz. In comparison, Traverso [74] reported 13% of 100 dog faeces positive to taeniid eggs in dogs from Guaqui in La Paz (near the Titicaca lake) and 20.4% (29/142) in dogs from Chucuito at the Titicaca lake in Peru. Gonzales [75] reported 9.1% (31/340) of canine faecal sample infected with taeniid eggs in Tupiza.

PLOS NEGLECTED TROPICAL DISEASES
First review of cystic echinococcosis in Bolivia

Cystic echinococcosis in livestock
No official documents from the government or scientific papers reporting CE in animals from Bolivia at a national level have been published. Table 4 summarises all the reports of CE in animals reviewed by us. Fig 1 shows the distribution of all the studies and their prevalences for different livestock species performed in Bolivia. The first mention of CE in animals from Bolivia is dated in 1910, in a book about the parasites found in the Bolivian Altiplano. Surprisingly, the disease was described in a horse and a donkey [76]. For this review, most data regarding CE in animals were acquired from theses, including 9 from the Faculty of Veterinary Medicine at the University Gabriel René Moreno in Santa Cruz de la Sierra; 1 thesis from the Universidad de El Alto, 1 from the Faculty of Agronomy at UMSA, and 1 from the Faculty of Veterinary Medicine at the University Tomas Frias ( The only study performed in Sucre included 699 cattle from 3 departments from which only 5 animals (0.7%) were infected with E. granulosus, including 1/37 from Santa Cruz (2.7%), 1/38 from Cochabamba (2.6%), and 3/607 from Chuquisaca (0.5%). At the same time, no infection was recorded in 17 animals from Potosí [83]. The only work investigating CE prevalence in sheep in the whole country described 38 out of 2,047 sheep (1.9%) positives from Batallas near the cities of La Paz and El Alto. According to the study, 45% of the sheep's cysts were unfertile [88]. Prevalences in cattle between 0% and 9.7% have been reported in Potosí [83,85]. Unfortunately, there is no published work referring to CE in goats in Bolivia. However, infected goats are found in the valleys in the south of the country.
Finally, the most recent description of CE in cattle from different Bolivia Departments was published as a thesis in 2001 by Reinoso [82], reporting a prevalence of 6.2% from 3,119 cattle. The highest prevalence was identified in animals slaughtered in official abattoirs in La Paz (8.3%). Some areas within the La Paz Department showed prevalences as high as 16% in Palcoco, 13.8% in Vilaque, and 7.8% in Batallas. Data from these theses show that the highest prevalences are at the northern Altiplano of the Department of La Paz, where the disease has been described in cattle, sheep, and pigs [41,82,88]. Unfortunately, only one of the theses included CE reports in sheep, which is the most important intermediate host for E. granulosus s.s. Finally, in the most recent survey of the prevalence of E. granulosus in animals in 2019, Calle [89]

Control of E. granulosus in Bolivia
An attempt to establish a national control programme for CE in Bolivia was proposed and mentioned in the First National Seminar for Control and Surveillance of Zoonoses in 1989 [91]. The plan was intended to last for 4 years; however, it was not implemented. A pilot study for diagnostic in humans and dogs was developed in Tupiza in collaboration with the "Southern cone sub regional initiative for control and surveillance of cystic echinococcosis" between 2009 and 2010. The results are shown in the "Ultrasound diagnosis, treatment, and patient follow-up" section of this review. Several recommendations and training in ultrasound were part of this pilot study [92]. Only a single dog deworming campaign was carried out [70]. Recently, the Ministry of Health established a guide for surveillance and prevention of E. granulosus and Fasciola hepatica [93].
On the other hand, the National Zoonosis Program of the Ministry of Health is limited to rabies vaccination of dogs, without activities against CE. Human CE notification is not compulsory, and the infected viscera from livestock are confiscated at the municipal abattoirs. Staff at the National Service of Agricultural Health and Food Safety (SENASAG) is notified, but not control measures are implemented. The sanitary inspection is limited to bovines in municipal abattoirs located in cities. The uncontrolled clandestine slaughter, mainly of pigs and sheep and South American camelids, persists even in communities close to large cities.

Epidemiological analysis of E. granulosus in Bolivia
Data reviewed here demonstrate that this zoonotic disease has been historically reported in 8 out of 9 Departments of this country except for Pando. The livestock population in Bolivia includes 9,304,572 bovines, 5,382,778 sheep, 2,941,827 pigs, 1,454,923 goats, and around 1 million South American camelids [94]. Large-scale intensive livestock rearing, specialised in cattle production, is restricted to the lowlands of the Departments of Beni, Santa Cruz, and Pando in the northeast of Bolivia. In contrast, in La Paz and other Departments, farmers have a small number of animals frequently mixing different species (bovines, ovine, pigs, and camelids). This happens because of the shortage of forage due to the Altiplano's bioclimatic conditions [95]. Livestock rearing is usually combined with other agricultural activities; supervising livestock is frequently the children's responsibility.
Considering the available data, we propose the analysis of the CE in 3 ecological levels in Bolivia: Altiplano (La Paz, Oruro, and Potosí), subtropical valleys (Cochabamba, Tarija, and Chuquisaca), and tropical lowlands from the Amazon (Beni, Pando, and Santa Cruz). In the case of the Altiplano, La Paz is the department that reported the majority of the cases and more retrospective studies on CE (see Tables 1 and 2). Unfortunately, the cases' actual origin is not available in several reports, and the majority of reports are from hospitals in big cities where surgeries were performed (passive detection). Cattle, sheep, and pigs are common in this area, principally in the ecoregion named "wet puna." The raising of South American camelids is scarce in this region. In our experience, the Northern Bolivian Altiplano at La Paz Department is one of the most endemic areas for CE in Bolivia. The transmission is low in certain valleys with few human cases reviewed here (Tables 1 and 2), while animal infection reached up to 37.5% in Tarija, for example (Table 4).
In contrast, 44 human cases have been reported in Chuquisaca, including the first autochthonous case from Bolivia; however, a low prevalence in livestock has been found in this department (Table 3). More precise information is available from Potosí's valleys. Only 1 epidemiological study involving several communities in Tupiza reveals the prevalence of CE between 2.1% and 13.3% in humans using ultrasound examination [62] and also the infection in dogs. In the case of the lowlands, livestock is found in large-scale intensive systems primarily cattle. Low CE prevalence has been described for this area for livestock, while no reports of CE in humans have been described in recent years. However, the identification of E. intermedius G7 in pigs from Santa Cruz [8] warrants further investigation in this Department. Previously, Echinococcus vogeli was identified in Santa Cruz infecting Cuniculus paca [96].

Conclusions
The available information on CE in Bolivia does not allow a comprehensive countrywide estimation of the disease's epidemiological situation and burden. However, data summarised here show that CE is present in 8 out of 9 Departments in Bolivia. CE became more commonly diagnosed in the last 50 years. The maximum levels of infection recorded in livestock in different species from Bolivia also show that the parasite is widely spread. Studies performed in dogs showed that the parasite is actively transmitted in areas near major population centres like La Paz and El Alto.
Furthermore, the recent detection of 3 different species of E. granulosus s.l. in Bolivia warrants further epidemiological investigations. Future baseline epidemiological data, including studies in dogs and intermediate hosts and a systematic collection of human cases, are necessary. Data acquired will form the basis for the discussion and implementation of national control programmes. Public health education will be essential due to the lack of knowledge of the disease in the Bolivian population. Promote the interest related to CE and the sensibility of the national and departmental authorities to assume influential and integral responsibilities in the context of "One Health" is an objective of this work. The management based on standardised protocols of the human CE in hospitals could clarify the disease's epidemiological situation in this country. Finally, further studies investigating other related species present in some South American countries including Bolivia, as E. vogeli or Echinococcus oligarthrus, are also indicated.