article: Rabies is a fatal viral disease that continues to threaten both human and animal health in endemic countries. The Lao People’s Democratic Republic (Lao PDR) is a rabies-endemic country in which dogs are the main reservoir and continue to present health risks for both human and animals throughout the country. Passive, laboratory–based rabies surveillance was performed for suspected cases of dog rabies in Vientiane Capital during 2010–2016 and eight additional provinces between 2015–2016 using the Direct Fluorescent Antibody Test (DFAT). There were 284 rabies positive cases from 415 dog samples submitted for diagnosis. 257 cases were from Vientiane Capital (2010–2016) and the remaining 27 cases were submitted during 2015–2016 from Champassak (16 cases), Vientiane Province (4 cases), Xieng Kuang (3 cases), Luang Prabang (2 cases), Saravan (1 case), Saisomboun (1 case) and Bokeo (1 case). There was a significant increase in rabies cases during the dry season (p = 0. 004) (November to April; i. e. , <100mm of rainfall per month). No significant differences were noted between age, sex, locality of rabies cases. The use of laboratory-based rabies surveillance is a useful method of monitoring rabies in Lao PDR and should be expanded to other provincial centers, particularly where there are active rabies control programs.
  Rabies is an acute viral encephalomyelitis caused by the rabies virus (genus Lyssavirus; family Rhabdoviridae) which can affect all warm-blooded animals including humans. Humans and animals may be infected with rabies virus via saliva through a bite or scratch [1] or butchering [2] of a rabid animal, and if left untreated it almost invariably leads to a fatal outcome. The World Health Organization estimates approximately 30,000 human deaths per year due to canine rabies in Asia out of a total 61,000 deaths per year due to rabies worldwide [3]. The Lao People’s Democratic Republic (Lao PDR) is a rabies-endemic country in which dogs are the main reservoir and continue to present health risks for both human and animals throughout the country [4,5]. Historical reports of rabies cases in Lao PDR have been recorded in humans since 1963 [6] and rabies cases in dogs have been reported since the late 1960s [7]. It has been estimated that an average of 8,528 dog bites are reported annually in Lao PDR [8] underlining the importance of rabies surveillance and control. More recently, studies have identified the increasing importance of dog rabies in Lao PDR [4], Cambodia [9], and southern China [10], examining diagnostic issues, case numbers, epidemiology and genetic relatedness of contemporary regional strains. Here we present results that 1) describe the number of dog rabies cases confirmed in Lao PDR from 2010–2016 using laboratory-based surveillance in Vientiane Capital, and including samples submitted from eight central, northern and southern provinces; 2) investigate aspects of dog rabies cases in Lao PDR including age, sex, location and season; 3) discuss opportunities for control of rabies in domestic animals using vaccination and identify constraints in the context of a low-resource environment.
  Diagnostic testing for the presence of rabies virus was performed at the National Animal Health Laboratory (NAHL), Ban Khunta, Sikhottabong district, Vientiane Capital, Lao PDR (see Fig 1 for location). Rabies-suspected animals originated from the nine districts of Vientiane Capital (Fig 1) during 2010–2016. From 2015–2016, samples were also submitted from Champassak, Luang Prabang, Saisomboun, Vientiane Province, Xieng Kuang, Bokeo, Saravan and Sekong provinces (Fig 2). Samples (carcass or head) presented for diagnosis came from animals that were strongly suspected of having rabies, exhibiting furious or comatose clinical signs, with the former often resulting in a human bite incident requiring rabies virus exclusion. In these circumstances, the bite-victim, and in many cases the animal owner, delivered the animals for urgent diagnostic testing. Rabies diagnostics on carcass material was performed following verbal agreement and consent of the animal owner and the bite victim so that compensation and treatment could be determined. Necropsy procedures were performed to allow sampling of the brain and collection of hippocampal tissue for diagnostic evaluation using the rabies virus direct fluorescent antibody test (DFAT) [11]. The DFAT requires a tissue impression or smear on a microscope slide stained with a FITC anti-rabies monoclonal antibody conjugate that is specific for rabies virus (Fujirebio Diagnostics Inc. , USA) which is visualized by fluorescent microscopy. All analysis was performed using STATA 14. 0 (StataCorp, College Station, TX USA). Descriptive statistics were used to characterise the number of rabies cases according to sex, age group, temporal and spatial distribution of rabies positive cases from 2010 to 2016. Significant differences (p<0. 05) in rabies positive cases was determined using Pearson’s Chi-Square (Chi2) analysis. Age was compared using the groupings of <3,3–12, >12–24, >24–36 and >36 months. Factors compared included the seasonal influence on rabies positive cases (wet season = >100mm of monthly rainfall i. e. May to October, compared to the dry season from November to April = <100mm of monthly rainfall [12]). Differences in the proportion of rabies positive samples from urban (Chanthabouly, Sisttanak, Sikhottabong and Saysettha) and rural (Naxaithong, Xaythany, Hatxaifong, Pakngum and Sangthong) districts of Vientiane Capital were also examined. Trends in the number of annual rabies cases and dog age were tested for significance (p<0. 05) using the ptrend command which calculates a Chi2 statistic for trend. Historical rabies case data from 1988–2011 was collected for comparative purposes. Data was derived from the same source being directly, or quoted as, provided by the Department of Livestock and Fisheries (DLF) of the Ministry of Agriculture, Lao PDR or by the Livestock and Veterinary Department (former title of DLF) from conference proceedings [13,14], and a peer reviewed publication [4] (Table 1). Data presented in Table 1 varies in terms of the number and provinces and species from where samples were submitted for diagnosis.
  During 2010–2016,415 dogs or specimens from dogs were submitted to NAHL for rabies diagnosis. The overall number of rabies positive cases was 284 (68. 4%). The numbers of samples submitted for rabies testing annually and the numbers of confirmed rabies cases is presented in Table 1. The median number of samples submitted annually was 57 (IQR: 51–73), with a range from 42 (2014) to 75 (2010). The median annual number of confirmed rabies cases was 36 (IQR: 32–53), range 29 (2014) to 57 (2015), and the annual proportion of rabies positive samples ranged from 56. 5% (2012) to 78. 1% (2015). There was no significant difference in rabies positivity rates (Chi2 = 10. 96; p = 0. 009) with no significant trend for the seven-year sampling period (Chi2 = 0. 497; p = 0. 481). The numbers of confirmed rabies cases by month from 2010–2016 are presented in Fig 3. During the entire 2010–2016 sampling period, the preponderance of dog rabies cases came from Vientiane Capital (90. 5%; 257/284) (Table 2) and of these majority were from Xanythany (37. 0%; 105/284), Sikhottabong (16. 2%; 46/284), Saysettha (15. 1%; 43/28) and Naxaithong (7. 8%; 22/284) districts. Fewer rabies cases were recorded from dogs in Hatxaifong (4. 9%), Sisattanak (4. 9%), Chantabouly (3. 9%) and Pakngum (0. 7%) districts, with no cases from Sangthong district (Table 2 and Fig 1). From 2015–2016, samples submitted from 8 provinces outside Vientiane Capital had at least one rabies case with the majority coming from the southern province of Champassak (6. 6%; 16/284) and the centrally located Vientiane Province (1. 4%; 4/284) (Table 2 and Fig 2). The number of rural rabies cases was higher than that of urban districts (i. e. , 166 cf 118) although this was not statistically significant (Chi2 = 0. 044; p = 0. 834). Focusing on rabies cases in Vientiane Capital, despite the larger number of samples from Xanythany, Sikhottabong, Saysettha and Naxaithong districts there was no statistical difference in the proportion of rabies cases between the districts (Chi2 = 12. 4; p = 0. 133) (Table 3). Fig 3 demonstrates annual frequency of rabies cases for the Vientiane Capital districts from 2010–2016. There was a significant seasonal difference in the number of rabies cases (Chi2 = 8. 32; p = 0. 004) with a higher number of cases and higher proportion of positive cases during the dry season (n = 173; 74. 3%) when compared to the wet season (n = 111; 61. 0%) (Table 3). There was a trend in the increase of monthly rabies cases from the start of the dry season (November) through to the start of the wet season (May) cases over the sampling period (Fig 4). Despite no significant monthly trend in rabies positive cases from January to December (Chi2 = 1. 38; p = 0. 240) there was a significant decrease in rabies cases in the 12-month period May to through to April (Chi2 = 8. 87; p = 0. 003) (Fig 4). There was no significant difference in the proportion of rabies positive cases between the sexes (males 62. 3%; Chi2 = 2. 12; p = 0. 145) or age groups (Chi2 = 4. 32; p = 0. 363) (Table 3). There was no significant trend in rabies cases with increasing age (Chi2 = 2. 17; p = 0. 141). The median age of rabies-positive animals (12 months; interquartile range (IQR) 4–24) was, however, more than double the median age of the negative animals (5 months (IQR: 3–12 months).
  Dog bite remains the main source of rabies transmission in the Lao PDR and dog vaccination remains the best method for rabies control. Dog owners can play an important role in rabies-control activities by ensuring that their dogs are vaccinated and restrained to reduce the opportunity for bites by rabies-infected animals. Governments and donors have an equally important role to play in providing a conducive environment and by facilitating increased awareness through mass media campaigns, effective delivery of vaccination and surveillance so that the situation can be actively monitored nationwide.
summary: Rabies is a viral disease that continues to threaten both human and animal health in endemic countries and almost always results in death of the infected individual. In most areas of Southeast Asia, rabies continues to be a major zoonosis with the main source of human cases being dog bites. In this study, we examined the brains of 415 rabies-suspected dogs from Vientiane Capital and some other provinces of Lao PDR during 2010–2016. Overall, rabies was confirmed in 284 cases. We found that there was a significant increase in rabies cases during the dry season (November to April) when monthly rainfall was less than <100mm. The laboratory-based surveillance of rabies in the domestic animal population is an important component of a rabies control program, however it should be used in conjunction with an effective vaccine program.
keywords: animal types
  medicine and health sciences
  pathology and laboratory medicine
  pathogens
  immunology
  tropical diseases
  microbiology
  vertebrates
  pets and companion animals
  dogs
  animals
  mammals
  viruses
  preventive medicine
  vaccines
  rabies
  rna viruses
  veterinary diagnostics
  neglected tropical diseases
  infectious disease control
  vaccination and immunization
  zoology
  veterinary science
  veterinary medicine
  rabies virus
  public and occupational health
  infectious diseases
  zoonoses
  medical microbiology
  microbial pathogens
  lyssavirus
  viral pathogens
  biology and life sciences
  viral diseases
  amniotes
  organisms
title: Rabies surveillance in dogs in Lao PDR from 2010-2016