3.1 Sampling Technique
Watercress was gathered along a variable length of each of the waterways depending on the
abundance of the watercress. Sampling commenced in the last week of March 2000 (except
for the golf club site) and continued for a total of five weeks. The reason for testing over a
five-week period was to assess variability in contaminant concentrations.
In the first week, five 300g watercress samples were collected from each of the sites for
individual microbiological analysis in order to be able to compare microbiological results
against the Ministry of Health’s “Microbiological Reference Criteria for Food 1995”. The
exception was Waiwhetu Stream where only one sample was collected each week as there
was insufficient watercress available. For the remaining four weeks, five samples (if
available) were taken from each site and combined by the laboratory into one sample for
analysis. In summary, a total of 9 watercress samples were analysed from each site except
from Waiwhetu where 5 samples were analysed.
Healthy looking watercress was selected randomly from the stream. The whole plant
excluding the roots was taken. A fresh pair of sterile disposable gloves was used to collect
each sample of watercress. Each sample was placed into an appropriately labelled sterile swirl
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bag and placed in a chilly bin with slika pads for overnight transportation to ESR for
microbiological and heavy metal analysis.
Growing waters were only sampled for microbiological contamination. However, testing of
growing waters (and potentially sediments) for evidence of heavy metal contamination would
be undertaken if watercress samples showed significant contamination. A 1L water sample
was taken from each of the sites simultaneously to watercress sampling. The water sample
was collected adjacent to where the watercress was collected. Each sample was collected in a
sterile bottle, appropriately labelled and placed in the chilly bin for overnight delivery to ESR.
3.2 Watercress Identification
To identify the species of watercress present at each site, seedheads and/or flowers (where
available) were sent to Rohan Wells, a Freshwater Ecologist at NIWA in Hamilton, for
identification:
• Papawai Stream: Rorippa microphylla
• Parkvale Stream: Rorippa microphylla & Rorippa nasturtium-aquaticum
• Manaia Drain: Rorippa microphylla & Rorippa nasturtium-aquaticum
• Opaki Stream: Rorippa nasturtium-aquaticum (Identified on the basis of flower
size only, not seed which is positive ID).
• Mazengarb Drain: Rorippa nasturtium-aquaticum
• Owhiro Stream: Rorippa microphylla
• Hulls Creek: Rorippa nasturtium-aquaticum
• Black Stream: Rorippa nasturtium-aquaticum & Rorippa microphylla
• Waiwhetu Stream: Rorippa microphylla
• Ohariu Stream: Rorippa nasturtium-aquaticum
• Golf Course Stream: (no flowers or seed capsules present to positively
identify the species).
Although two species were identified, the morphology of both is similar and therefore a
significant variation in contaminant concentrations between the two species of watercress
would not be expected.
3.3 Microbiological Analysis
3.3.1 Summary
Microbiological testing of the watercress included bacterial counts for presumptive coliforms,
faecal coliforms, Escherichia coli (E. coli) and presence/absence tests for Campylobacter
species. Microbiological testing of the growing waters included bacterial counts for total
coliforms, E. coli and presence/absence tests for Campylobacter species.
Of all the coliforms, E. coli is the most specific indicator of faecal contamination readily
available (MfE, 1999b). The public health significance often depends on their origin, but
generally, E. coli levels are taken as an indicator of the degree of faecal pollution, and the
potential for enteric pathogens to be present.
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E. coli comes from the family of bacteria known as Enterobacteriaceae and is the most
common bacteria of this group (MfE, 1999b). It is nearly always found in the gut of humans
and animals, usually in high numbers. It can survive for up to four to six weeks in fresh water
especially when shaded from sunlight and is a definite indication of recent faecal
contamination (MfE, 1999b).
E. coli is the preferred indicator organism for fresh waters and was therefore chosen as the
most appropriate indicator organism for testing watercress and growing waters. The
probability of E. coli multiplying in water is very small so the number detected relates to the
original level of faecal contamination (MfE, 1999b).
There are many different types, or strains, of E. coli. Many are harmless to humans, but some
pathogenic strains of E. coli are very serious e.g. E. coli 0157, because of the symptoms they
cause and the extremely low dosage required to cause these symptoms. E. coli 0157, which is
transmitted from ruminants, enters the food chain primarily by contaminated foods (Ball and
Till, 1998). The significance of waterborne E. coli 0157 in New Zealand is not known (Ball
and Till, 1998); however, E. coli 0157:H7 was detected in two of 531 faecal specimens in a
survey of healthy dairy cattle (Buncic and Avery, 1997).
Campylobacter is a common food and water-borne pathogen and is the most frequently
notified food-borne disease in New Zealand (Ministry of Health, 1998). There is good
evidence to implicate contaminated drinking water in several Campylobacteriosis outbreaks
in rural areas of New Zealand (Ball and Till, 1998). 3.3.2 Laboratory Analysis
Microbiological analyses were undertaken by the ESR Christchurch Public Health
Laboratory, an accredited IANZ laboratory.
Each sample, or composite of five samples, of watercress was tested for the following
bacteria:
• E. coli (presumptive coliforms & faecal coliforms were also measured as part of the
method for measuring E. coli).
• Campylobacter species.
Water samples were tested for:
• E. coli (total coliforms were also measured as part of the method for measuring E. coli).
• Campylobacter species.
E. coli:
A 50 g portion of each watercress sample was weighed into a sterile Stomacher bag and
homogenised with 100 ml of sterile 0.1% Peptone water. Serial dilutions from 10-1 to 10-3
were prepared and 5 tube MPNs were set up using Laurel sulphate broth. These were
incubated at 35o
C for a total of 48 hours. Positive tubes were subcultured to EC Broth and
incubated for 24 hours at 44.5o
C. Faecal coliforms from these tubes were further confirmed
to E. coli using EMB agar plates, BGBB and Indole production (limits of detection, <1.8,
>2400MPN/g).
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Water samples from each watercress growing site were tested for total coliforms and E. coli
using “Colilert” MPN (limits of detection, <1, >2400 MPN/100ml).
Campylobacter spp:
A 10-g portion of watercress was weighed into 90 ml of “Exeter Broth” and incubated at 42o
C
in a reduced oxygen atmosphere (produced by using a “Campy-Gen” envelope in a Gas-Pak
jar). After 48 ± 2 hours incubation the broth was subcultured onto an “Exeter agar” plate and
incubated as for the broth. Suspicious colonies isolated on this plate were confirmed as being
Campylobacter spp by Gram stain, catalase and oxidase tests.
A 1 litre volume of water from each watercress growing site was filtered through a 45 µ
membrane filter which was then placed in 90 ml of “Exeter Broth”. This broth was then
treated as for the watercress sample.
write the main points for the above information given
1. Watercress sampling was conducted over a five-week period to assess variability in contaminant concentrations.
2. Healthy watercress samples were randomly selected from each site for analysis.
3. Watercress species were identified at each site by sending seedheads and flowers to an expert.
4. Microbiological testing of watercress and growing waters included bacterial counts for E. coli, fecal coliforms, and total coliforms, as well as presence/absence tests for Campylobacter species.
5. E. coli is the preferred indicator organism for testing water quality as it indicates recent fecal contamination.
6. Campylobacter is a common food and waterborne pathogen in New Zealand.
7. Microbiological analyses were conducted by an accredited IANZ laboratory.
8. Laboratory analysis methods included microbiological testing for E. coli and Campylobacter species in watercress and water samples.
9. Confirmation tests were conducted for E. coli using EMB agar plates, BGBB, and Indole production.
10. Campylobacter species were confirmed through Gram stain, catalase, and oxidase tests.