Campylobacter is a Gram-negative, thermophilic and microaerophilic bacterium that causes approximately 5-14% of diarrheal illness worldwide (Nachamkin et al., 2008; WHO, 2004). Most of the identified Campylobacter species can cause human campylobacteriosis but, of these, Campylobacter jejuni and C. coli are the major causes of gastroenteritis, accounting for 95% of all reported human campylobacteriosis cases (WHO, 2004). This organism naturally inhabits the intestines of warm-blooded animals including poultry, wild birds and ruminants (Stanley and Jones, 2003). Several epidemiological studies have shown these animals to be potential reservoirs for human infections (Stanley and Jones, 2003; Wong et al., 2006; French et al., 2009). These studies also indicate that animal hosts are the main sources of food and water contamination, whilst the food chain route has been shown to be the predominant infection pathway for human campylobacteriosis, particularly via poultry meat (Corry and Atabay, 2001; Woodward et al., 2005). Sporadic cases of Campylobacter infection have been most commonly linked to food (ESR, 2011b; Samuel et al., 2004), whereas outbreaks of campylobacteriosis have been associated with drinking contaminated water, the accidental ingestion of water during recreational activities, and the consumption of poultry meat and raw milk (Evans et al., 1996; Frost et al., 2002; Schönberg-Norio et al., 2004). Hence, each potential exposure pathway needs to be studied in detail in order to increase our understanding of Campylobacter transmission dynamics and thus inform the design of effective, country-specific, prevention programmes.
Campylobacteriosis is the most common zoonotic bacterial enteric disease in New Zealand, and over the last decade, this country has persistently reported one of the highest campylobacteriosis rates among developed countries (Wilson, 2005; Baker et
60
al., 2007b; Muellner et al., 2011). From 1980 (the first year of mandatory notification of cases) to 2006, the annual number of reported cases of campylobacteriosis increased steadily from a few hundred to 15,873 (Baker et al., 2007a; ESR, 2007; ESR, 2011b). These cases accounted for 70% of all the notified enteric diseases in New Zealand and resulted in an estimated cost of NZD 75 million per year (Scott et al., 2000; NZFSA, 2010). Around 25% of campylobacteriosis cases incurred a further disease burden due to sequelae such as Guillain-Barré syndrome with an average rate of 2.32 hospitalisations per 100 000 population per year (Baker et al., 2012; Muellner et al., 2009). An investigation into the sources of human campylobacteriosis cases identified poultry as a major source of infection (Wilson, 2005). Subsequently, in early 2007, the three major poultry suppliers implemented voluntary and regulatory interventions to reduce poultry carcasss contamination. In the two years subsequent to the interventions, there was a 59% reduction in reported human campylobacteriosis cases from 383 cases in 2006 to 157 cases per 100,000 population in 2008) (Sears et al., 2011). Nevertheless, the campylobacteriosis rate in New Zealand remains among the highest of the industrialised countries, indicating the need to understand the role of other sources of infection, including water and ruminants.
In New Zealand, thermophilic Campylobacter spp. have been isolated from river water, streams, lakes, ponds, runoff water, and drinking water (Koenraad et al., 1997; Till et al., 2008). Savill and co-workers (Savill et al., 2001) found that 60% of 30 recreational water sites were positive, with counts of Campylobacter reaching 11 most probable number (MPN1) 100 mL-1. Between December 1998 and February 2000, a large-scale
survey was conducted in 25 freshwater recreational and drinking water supply sites distributed throughout New Zealand (Till et al., 2008). This study showed that 60% of the 725 samples were positive for Campylobacter spp. with 48% of the positive samples identified as C. jejuni. Till et al. (2008) also estimated that 5% of campylobacteriosis cases could be attributed to recreational water. This freshwater survey and a quantitative risk assessment led to develop a new national recreational freshwater water quality guideline in 2003. However, applying these guidelines to recreational rivers is complicated because many factors, including the rate of river flow, land use, animal access to the waterways and surface runoff, could influence the bacterial risks to human health.
1 Most probable number (MPN) is the method to estimate the concentration of viable microorganisms in a sample by
means of replicate liquid broth growth in ten-fold dilutions and is particularly useful with samples that contain low concentrations of organisms (<100/g) (Blodgette, 2010)
61
Campylobacter spp. only replicate in animals, and although they are frequently recovered from the environmental water, they have not been demonstrated to multiply outside the host (WHO, 2004). The growth of C. jejuni and C. coli requires both thermophilic (>30 oC) and microaerophilic (<15% of O2 and CO2) atmospheric
conditions (WHO, 2004). However, it has been suggested that Campylobacter spp. may persist in the environment by entering a viable but non-culturable (VBNC) state, and/or by forming monospecies biofilm or colonising pre-existing biofilm (Chaisowwong et al., 2011; Reuter et al., 2010). Even so, ambient temperatures, high oxygen concentrations, UV radiation, and desiccation may decrease the survival of
Campylobacter in the environment (Rollins et al., 1986; WHO, 2004; Inglis et al., 2010). Therefore, the presence of Campylobacter in water most likely indicates recent faecal contamination from either a point source (e.g. meat plant effluent) or nonpoint sources (e.g. agricultural runoff).
The subtyping of C. jejuni has been used successfully to identify the sources of human infections. Several methodologies, including serotyping and PFGE have been developed for subtyping of C. jejuni (Lorenz et al., 1998; Nielsen et al., 2000). However, multilocus sequence typing has good discriminatory power and better reproducibility than other typing methods (Dingle et al., 2001). Further, the use of MLST for typing C. jejuni has also provided important insights into the population genetics of this organism (Dingle et al., 2001; 2005) and has helped to increase the understanding of transmission pathways of human campylobacteriosis (Wilson et al., 2008). Source attribution modelling has also been used in human campylobacteriosis cases in the Manawatu, New Zealand. Prior to intervention in the poultry industry, an estimated 70% of human cases were attributable to poultry sources. Subsequently, there has been a decrease in the proportion of human cases attributed to poultry and an increase in the proportion of cases attributable to ruminant sources, which suggest that livestock are an important source of infection, particularly in rural areas (Dingle et al., 2005; Muellner et al., 2011).
Previous investigations into Campylobacter in rivers showed evidence of seasonal differences in retrieving Campylobacter from water samples (Jones, 2001; Obiri-Danso et al., 2001). In addition, diverse sequence types of C. jejuni have been isolated from rivers, and the majority of sequence types identified as those associated with wild birds (Obiri-Danso et al., 2001). Here, we report the findings of a longitudinal study of
Campylobacter in six high-use recreational rivers in the Manawatu region from 2005 to 2009. The primary objectives of this study were:
62
1) to assess the proportions of Campylobacter spp. and C. jejuni positive water samples from each of the study sites, and
2) to determine the potential associations between the presence of both Campylobacter
spp. and ruminant-associated C. jejuni and explanatory variables such as month and site of sample collection, and river flow rates.
In addition, the population genetic structure of C. jejuni was assessed to explore possible animal sources of river-borne isolates, and the potential of the strains present to cause human infection.