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Microbial Assessment Of Plastic Bottles Reused For Packaging Food Products In

Ghana

Sika Abrokwah, Bernard Ekumah, Francis Abrokwah

PII: S0956-7135(19)30545-6
DOI: https://doi.org/10.1016/j.foodcont.2019.106956
Reference: JFCO 106956

To appear in: Food Control

Received Date: 25 July 2019

Accepted Date: 15 October 2019

Please cite this article as: Sika Abrokwah, Bernard Ekumah, Francis Abrokwah, Microbial
Assessment Of Plastic Bottles Reused For Packaging Food Products In Ghana, Food Control
(2019), https://doi.org/10.1016/j.foodcont.2019.106956

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© 2019 Published by Elsevier.

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MICROBIAL ASSESSMENT OF PLASTIC BOTTLES REUSED FOR PACKAGING FOOD

PRODUCTS IN GHANA

Sika Abrokwah1*, Bernard Ekumah2 and Francis Abrokwah3

1. Centre for Coastal Management, University of Cape Coast, Cape Coast, Ghana
2. Department of Environmental Science, University of Cape Coast, Cape Coast, Ghana
3. Department of Biochemistry, University of Cape Coast, Cape Coast, Ghana

* E-mail of the corresponding author: sika.abrokwah@ucc.edu.gh

Abstract
Patronage of bottled water in Ghana has increased over the years as a more hygienic source of drinking
water. This shift has however resulted in increased generation of waste bottles without any formal regulated
management system to handle them sustainably. This has encouraged an informal system which involves
the retrieval and reuse of the bottles by market women and small scale local beverage producers for the sale
of products such as palm oil, mashed kenkey and hibiscus tea (“sobolo”). The reuse of these bottles for food
products for public consumption without any regulated means of sanitizing them is an issue of public health
concern. This study therefore sought to determine the level of microbial contamination in used bottles sold
in the three major markets (Makola, Kaneshie and Madina) in the capital city of Ghana, Accra to ascertain
their safety for use for food products. The results showed presence of Total Coliforms and Faecal Coliforms
in more than 60.0% and 40.0% of the bottles respectively. The bottles washed by the vendors recorded
higher presence of Total Coliforms (65.3%) and Faecal Coliforms (57.3%) than the unwashed ones (49.3%)
and (36.0%) respectively. This could be attributed to poor water quality used in washing the bottles, cross
contamination from washing equipment, poor washing technique/procedure and improper storage. The
continued reuse of bottles for food products therefore presents a serious public health problem. There is the
urgent need for strict enforcement of food safety regulations in Ghana to ensure that packaging materials
used by food vendors as well as the mode of treating these materials meet standards that safeguard public
health.

Keywords: Plastic Bottles, Food Packaging, Reuse, Contamination, Coliform, Treatment, Public Health

1. Introduction
In Ghana, access to safe drinking water and sanitation has been a challenge over the years. According to
the Ghana Demographic and Housing Survey (GDHS) 2014, thirty percent (30%) of households in Ghana
rely on sachet and bottled water as their main source of drinking water. The sachet water industry however
has come under attack in recent times over doubts of water quality as well as the quantum of waste the
sector generates. Several studies have reported microbial contamination of some sachet water (Obiri-Danso
et al., 2003; Dodoo et al., 2006; Ampofo et al., 2007; Kwakye-Nuako et al., 2007; Okioga, 2007; Addo et
al., 2009). Bottled water which used to be the preserve of the rich due to higher price (Stoler et al., 2012)
is increasingly gaining higher patronage from the general public as a result of growing mistrust of the
quality of sachet water. A total of thirteen (13) water bottling companies were certified by the Ghana
Standards Authority in December 2018 (Ghana Standards Authority, 2019). Although this shift may be
positive in providing a more reliable source of drinking water, it has increased the generation of used plastic
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bottles without any formal regulated management system to handle them sustainably. Even though used
plastic bottles are among the most readily recycled plastic products, currently, the rate of recycling stands
as low as 10% in some countries (Organization for Economic Co-operation and Development (OECD),
2018). The rest end up in landfills, incinerators and in the oceans. In the United States (US), the largest
producer of this waste, only about 20% of the bottles are recycled (United States Environmental Protection
Agency (US EPA), 2011).

In Ghana, there is very little published information on the fate of used plastic bottles generated in the
country. According to Fobil and Hogarh (2006), these bottles are traditionally sought after for reuse in
homes, by traders and local beverage manufacturers. This has encouraged an informal unregulated
management system which involves the retrieval and reuse of the bottles by market women and small scale
local beverage producers. The bottles are used for the sale of products such as palm oil and local beverages
such as mashed kenkey and hibiscus tea (sobolo). The informal sector that relies on these used bottles,
mainly source them from waste pickers who collect them either directly at the source of generation (e.g.
homes, social gatherings) or retrieve them from final disposal sites.

The reuse of these bottles for food products for public consumption without any regulated means of
sanitizing them is an issue of public health concern. Reusing plastic bottles which have not been adequately
washed and dried can result in the spread of bacteria from the hands and mouths (Booyens, 2012).
According to Reynolds (2005), improper maintenance of plastic beverage bottles can lead to contamination
with human faecal bacteria, or other harmful microbes. A study carried out by Obeng et. al. in 2014 on
microbial contamination in hibiscus tea packaged in both used and unused plastic bottles observed that
samples in used bottles were more contaminated (mean viable count 5.27 ± 0.87 x 102 CFU/ml) than
samples in unused plastic bottles (mean viable count 4.39 ± 0.74 x 102CFU/ml). Globally, food safety has
become an issue of concern due to the negative impact foodborne diseases on the socio-economic
development of nations. The WHO estimated that in 2010 alone, 230,000 deaths occurred worldwide
caused by foodborne diarrhoeal agents. In the developing world, challenges of unsafe sources of water for
food preparation, lack of proper pre- and post-processing food storage and low compliance and enforcement
of regulations have made foodborne diseases a typical occurrence affecting particularly children under the
age of five (WHO, 2015).

This study therefore sought to determine the level of contamination of Total and Faecal Coliforms in used
plastic bottles sold in three major markets (Makola, Kaneshie and Madina) in Accra, the capital city of
Ghana to ascertain their safety for reuse for food products.

2. Methodology

2.1 Study Area

The study was carried out at Makola, Kaneshie and Madina markets located in Accra, the capital city of
Ghana. These markets were selected because they are major markets from which vendors and street hawkers
of palm oil and local beverages source their used plastic bottles from.

2.2 Sample Collection

A total of 200 Polyethylene terephthalate (PET) bottles were collected for analysis made up of 25 unwashed
used bottles from each Market, 25 washed used bottles from each market, 25 used bottles from 5 homes in
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Accra and 25 new bottles as control. The Ghana Standards Authority in 2017 specified that the appropriate
number of samples to be obtained for each lot of packaged water considered for water quality analysis
should range from 15 to 24. As there is currently no standard for microbiological quality analysis of
packaging materials in Ghana, the standard for water quality analysis was used in this study. This
requirement therefore informed the choice of the number of bottles for each category of bottles tested.

2.3 Microbial Analyses

2.3.1 Agar preparation and Coliform counts
Total Coliform group includes both Faecal and environmental species are indicators of microbiological
presence in water. Most coliform bacteria do not cause disease but their presence is a cause for concern
because of the potential for other disease causing strains of bacteria to also be present. Faecal coliforms
are a subcomponent of this larger group of coliform bacteria. Faecal coliform are indicator organisms for
faecal contamination and potential presence of enteric pathogens (Baron, 1996). The samples were
analysed for both Total Coliforms and Faecal Coliforms as indicators of microbiological contamination.

Total Coliforms were estimated using the pour plate method with some slight modifications. Briefly, the
inner portion of each bottle was washed with 9ml of 0.85% physiological saline, these served as inoculum.
A total volume of 1ml of each inoculum was measured with a sterile 1ml disposable pipette and aseptically
dispensed into a well-labelled Petri dish. A total volume of 10ml of double strength melted Violet – Red –
Blue (VRB) Agar was added to each Petri dish. Each Petri dish was securely covered and the agar allowed
to solidify. Each Petri dish was turned upside down and incubated for 18 – 24 hours in the Laminar Flow
Hood at a temperature of 35°C. The total number of colonies were counted and recorded in CFU/ml. All
processes were carried out within the Laminar Flow Hood to prevent external contamination. Similar
method was used to estimate the Faecal Coliforms, this time using MacConkey Agar as the medium.

2.3.2 Quality Assurance

In order to ensure that the results of analysis obtained were accurate, quality assurance measures were
observed as follows:
i. Samples were analysed based on WHO’s Analytical Methods for Microbiological Water Quality
Testing
ii. All the glassware were thoroughly cleansed with appropriate detergent and rinsed with distilled
water and autoclaved. Pair of scissors, petri dishes, beakers, automatic pipette, agar, distilled water
were all autoclaved.
iii. Disinfection of the work area (the Laminar Flow Hood) and hands with 70% ethanol.

2.3.3 Data Analyses

All statistical analyses were carried out in Stata 13. Descriptive statistics was done covering mean viable
count, frequency and percentage occurrence of microbial load. The study employed confidence interval of
95% and statistical significance value of 0.05.

Negative log-log regression was fitted to the data to evaluate presence or absence of Faecal Coliforms in
the used bottles. Negative log-log regression was selected because bottle without Faecal Coliform were
more than the bottles that recorded the presence of Faecal Coliform. The dependent variable was
represented as dichotomous variable, with ‘1’ representing presence of Faecal Coliform and ‘0’
representing absence of Faecal Coliform. The independent variables were market, treatment and
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combination of market and treatment. Madina market was selected as reference for market because Madina
is relatively smaller as compared to the other two markets and also, it is located at the periphery of Accra.
With regards to treatment, washed bottle was chosen as the reference because ideally, cleaned used bottle
is expected to be less contaminated as compared to unwashed used bottle. The presence of coliform was
reported using exponentiated coefficient.

Kruskal-Wallis Equality of Populations Rank Test was conducted to find out if microbial load in the used
plastic bottles differs among markets and between treatments. A non-parametric test was preferred for this
analysis because microbial load is a count variable.

3. Results
Out of a total of 75 washed and 75 unwashed bottles tested from the three major markets in Accra, the
washed bottles generally recorded higher presence of microbiological organisms than the unwashed bottles
(see Table 1 and 2). The washed bottles had a mean viable count of 53.0CFU/ml for Total Coliform as
compared to the unwashed bottles with a mean viable count of 46.1CFU/ml. The bottles picked directly
from homes had a mean viable count of Total Coliforms of 0.8CFU/ml while the new bottles had 0CFU/ml
(Table 1). For Faecal Coliforms, the washed bottles again recorded the highest mean viable count of
29.3CFU/ml followed by the unwashed bottles at 16.3CFU/ml. The bottles picked from homes had a mean
viable count of 0.5CFU/ml and the new bottles 0CFU/ml (Table 2). The percentage occurrence of both
Total Coliforms and Faecal Coliforms in the washed bottles was 65.3% and 49.3% respectively and in the
unwashed bottles was 57.3% and 37.0% respectively. The bottles from home had 4.0% contamination for
both Total and Faecal Coliforms and the new bottles had 0% contaminated (Table 1 and 2).

Table 1 (Here)

Table 2 (Here)

3.1 Percentage Occurrence of Total Coliforms and Faecal Coliforms

Comparing the percentage occurrence of the Total Coliforms on washed bottles from the three markets,
Madina recorded the highest level of contamination of Total Coliforms (72.0%) with Makola and Kaneshie
recording (64.0%) and (60.0%) respectively (Fig 1). For the Faecal Coliforms, it was observed that bottles
from Madina again recorded the highest (64.0%) level of contamination on the washed bottles while 52.0%
and 28.0% were recorded in Makola and Kaneshie respectively. The details of percentage occurrence of
Total Coliforms and Faecal Coliforms are provided in Figure 1 and 2 respectively.

Figure 1(Here)

Figure 2(Here)

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3.2 Mean Viable Counts of Total and Faecal Coliform

Figure 3 and 4 show the mean Total Coliform count and the mean Faecal Coliform count respectively
detected in the washed and unwashed bottles from the three markets. Madina recorded the highest mean
Total Coliform count in both washed and unwashed bottles with Kaneshie recording the lowest (Fig. 3).
Makola recorded the highest mean Faecal Coliform count in the washed bottles and Madina had the lowest
mean Faecal Coliform count for the unwashed bottles.

Figure 3 (Here)

Figure 4 (Here)

3.3 Kruskal-Wallis Equality of Populations Rank test

Kruskal-Wallis Equality of Populations Rank test was used to test for the significance of difference between
the microbial load on the washed bottles and unwashed bottles as well as between the microbial load on the
bottles from the different markets. The results as presented in Table 3, showed that there was no significant
difference between the washed and unwashed bottles for both Total Coliforms and Faecal Coliforms.
However there was a significant difference between Total Coliform on bottles from the markets.

Table 3 (Here)

3.4 Statistical Models

Negative log-log regression of Faecal Coliform presence (Table 4) showed that bottles (washed and
unwashed) from Kaneshie Market were 65.0% less likely to record presence of Faecal Coliforms as
compared to bottles from Madina. Also, unwashed bottles are 60.8% less likely to record Faecal Coliforms
as compared to washed bottles.

Table 4 (Here)

4. Discussion

4.1 Presence of Total and Faecal Coliforms

As can be seen from the results, of all the bottles tested only the new unused bottles (control) showed no
presence of Total or Faecal coliforms. Both washed and unwashed bottles showed presence of both Total
and Faecal coliforms. All the vendors, with the exception of one vendor in Kaneshie, source their bottles
from waste pickers. That one vendor in Kaneshie indicated that she sources used bottles directly from users
such as family and friends. This may have accounted for relatively lower contamination levels observed in
bottles from Kaneshie. As shown earlier, used bottles taken directly from home had only 1 representing
4.0% of the bottles contaminated with coliforms (Table 1). Further, the results showed that even though
both washed and unwashed bottles generally showed some presence of Total and Faecal coliforms, the
washed bottles recorded a higher percentage occurrence of presence of coliforms than the unwashed. The

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results also show that unwashed bottles were 61.0% less likely to record Faecal Coliforms as compared to
washed bottles (Table 4). This pattern is interesting because the washing of the bottles primarily is to
sanitize them and remove contamination, however, as shown in the results, this process did not improve the
quality of the bottles but rather increased the chances of contamination. This pattern is also noteworthy
because if the washing and associated processes of the used bottles appeared to increase the presence of the
coliforms then washing and/or handling of new packaging materials or other food products such as fruits
and vegetables by market women or street vendors could introduce coliforms and other bacteria which
would render even new packaging materials or food products unwholesome for consumption. The increase
in contamination levels of washed bottles may be attributed to the quality of water used in washing, the
contamination level of the washing equipment, the washing procedure and/or storage.

Quality of Washing Water: In the Accra Metropolis, the Ghana Water Company (GWCL) produces most
of the water used but only about 45.0% of the population has access to this supply directly in their homes
or yards (Van-Rooijen et. al., 2008). The rest depend on private and community service providers who
either also get their supply from the GWCL, through tanker services or wells and boreholes (Adank et al,
2011). Thus, with a situation like this, it is difficult to determine where these bottle vendors access the
water used in washing the bottles. According to the Ghana Multiple Indicator Cluster Survey conducted in
2017 and 2018, on the average forty-eight percent (48.0%) of drinking water sources in Ghana showed
presence of Escherichia Coli (Ghana Statistical Service, 2019).

Washing equipment: The washing equipment which include the sponge/foam and buckets/washing bowls
may already be contaminated with coliforms and may transfer the contamination to the bottles. The washing
equipment used by the vendors of the bottles do not undergo any form of prior disinfection before usage.
Hazard Critical Control Points (HACCP) Standard Operating Procedures require that to ensure food
hygiene and safety, cleaning equipment should be sanitized and allowed to air dry between uses or after
four hours if in constant use (Bonne et. al., 2005).

Washing technique/procedure: How frequently the washing water is changed and how many rinses the
bottle is taken through may also increase contamination. The WHO Code of Hygienic Practice for
Bottled/Packaged Drinking Waters (2001) states that “reused containers should be washed and disinfected
in an appropriate system and positioned within the processing plant so as to minimize post-sanitizing
contamination prior to filling and sealing.” Hazard Critical Control Points (HACCP) standards require
that water used for washing should be clean, free of grease and food particles and also washing be done in
a three separate compartment that is, washing compartment, rinsing compartment and sanitizing
compartment each with defined temperatures (Bonne et. al., 2005). From observation in Madina market
where the washing of the bottles was done on-site, water used for both washing and rinsing the bottles are
reused several times before changed, if at all. This could lead to cross contamination and may have
accounted for the high contamination observed in washed bottles at this location (Fig. 1 and 2).

Storage: Finally, from observation, washed bottles are stored in sacks without drying awaiting purchase.
The moisture provides an ideal environment for bacterial re-growth and multiplication. Bacteria need water
to dissolve the food they use for energy and growth. Water allows the food to get into the cells, is used for
the many chemical reactions necessary for life and growth, and allows waste products to escape. In ideal
conditions (i.e. in moist foods at 37°C) bacteria will grow and multiply several times. After 6 hours, in
ideal conditions, one bacterial cell could become 131,072 bacteria (Wilmcow, 2012).

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4.2 Health Implications of Used Plastic bottles for Food products

The presence of total coliform is an indication that other more serious health threatening bacteria may be
present. The presence of Faecal coliforms however confirms the presence of such serious health threatening
bacteria. It is an indication that the bottles have been contaminated by faecal matter which could have been
from the garbage from which the bottles were picked or the water used to wash the bottles. Consumption
of food containing Faecal coliforms could cause dysentery, typhoid fever, hepatitis A, viral and bacterial
gastroenteritis (Nkere et. al., 2011). The usage of these bottles for packaging food for direct consumption
puts consumers at risk of contracting any of these diseases which may lead to not only economic losses due
to increased medical bills and loss of productive working hours but even death. According to the Food and
Drugs Authority in 2006, 90692 persons in Ghana died from food and personal hygiene related diseases.

The Codex Alimentarius Commission’s General Principles of Food Hygiene (2003) requires that
“packaging materials must be non-toxic and not pose a threat to the safety and suitability of food under the
specified conditions of storage and use. Where appropriate, reusable packaging should be suitably durable,
easy to clean and, where necessary, disinfect.” The used plastic bottles do not meet any of these criteria
but continue to be reused all over Ghana for packaging food without recourse to food safety standards. The
Public Health Act 2012 (Act 851) states that it is an offence to sell, prepare, package, convey, store or
display for sale, food under insanitary conditions to the public and it is also an offence to sell food that is
unfit for human consumption. Clearly from the contamination levels observed, these used bottles are
insanitary and would render any food packaged or stored in them unfit for human consumption. The Act
also spells out clear sanctions for persons found contravening these regulations. There is the need to
educate the players in the informal food processing sector such as market women, street vendors and
consumers on existing standards and procedures for choosing and handling food and packaging materials
as well as the legal implications of non-compliance with the standards. The regulations and punitive
measures detailed in Act 851 should be strictly enforced by the regulatory authorities. Without education
and enforcement of these standards, new bottles, other new packaging materials and food products may
also be contaminated during handling by the food processors.

5. Conclusion
The presence of Total and Faecal Coliforms was observed on each lot of used bottles sampled. Washed
bottles had higher percentage presence of both Total and Faecal Coliforms than the unwashed. Washing
appeared to increase microbial contamination on the bottles in all the markets. The used bottles are reused
for food products including drinks that are consumed directly without any further processing, the presence
of coliforms is an indication that this practice is unacceptable and presents a serious health risk to
consumers. There is the need for public education and strict enforcement of regulations and standards
governing the food processing sector. The results of this study also calls for a holistic waste management
policy in Ghana that will reduce the generation of used plastic bottles, eliminate their indiscriminate
disposal and improve collection for recycling. This will provide income generating opportunities for
Ghanaians without compromising the health of the general public.

6. Acknowledgements
The authors express their profound gratitude to Prof. Martin Oteng-Ababio and Dr. Ted Y. Annang for
comments on earlier versions of this paper.
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7. Funding
This research did not receive any specific grant from funding agencies in the public, commercial, or not-
for-profit sectors.

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Conflict of Interest and Authorship Conformation Form

Please check the following as appropriate:

o All authors have participated in (a) conception and design, or analysis and
interpretation of the data; (b) drafting the article or revising it critically for
important intellectual content; and (c) approval of the final version.

o This manuscript has not been submitted to, nor is under review at, another
journal or other publishing venue.

o The authors have no affiliation with any organization with a direct or indirect
financial interest in the subject matter discussed in the manuscript

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indirect financial interest in the subject matter discussed in the manuscript:

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Percentage Occurrence of Total Coliforms

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60 64 64
60
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48
40
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30

20

10

0
Washed Unwashed Washed Unwashed Washed Unwashed
Madina Makola Kaneshie
Figure 1: Percentage Occurrence of Total Coliforms in Washed and Unwashed bottles from the Madina,
Makola and Kaneshie Markets

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Pecentage Occurrence of Faecal Coliforms

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64
50
52
40
40
36
30
28
20 28

10

0
Washed Unwashed Washed Unwashed Washed Unwashed
Madina Makola Kaneshie
Figure 2: Percentage Occurrence of Faecal Coliforms in Washed and Unwashed bottles from the Madina,
Makola and Kaneshie.
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Figure 3: Mean Total Coliform in Washed and Unwashed bottles from Madina, Makola and Kaneshie
markets

Figure 4: Mean Total Coliform in Washed and Unwashed bottles from Makola, Madina and Kaneshie
markets
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Highlights:
 Used plastic bottles are reused for food products in Ghana
 These bottles showed presence of both Total and Faecal Coliforms
 The washed bottles showed higher presence of Coliforms than the unwashed bottles
 The reuse of plastic bottles for food products is unacceptable
 There is the need for regulations for packaging materials for food products
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Table 1: Total Coliforms in Washed and Unwashed Bottles

Bottle Type
New bottles
Parameter Washed Unwashed Home (Control)
Mean Viable Count (CFU/ml) 53.0 46.1 0.8 0
Standard Deviation 68.2 74.8 3.8
Percentage occurrence (%)
Presence of Total Coliforms 65.3 57.3 4.0 0
GSA/WHO Standard 0 0 0 0

Table 2: Faecal Coliform in Washed and Unwashed Bottles

Bottle Type
New bottles
Parameter Washed Unwashed Home (Control)
Mean Viable Count (CFU/ml) 29.3 16.3 0.5 0
Standard Deviation 45.9 34.8 2.4
Percentage occurrence (%)
Presence of Faecal Coliform 49.3 36.0 4.0 0
GSA/WHO Standard 0 0 0 0

Table 3: Kruskal-Wallis Equality of Populations Rank Test

Outcome/Response Factor/Group df χ2 P value
Market 2 8.986 0.0156
Total Coliform Treatment (washing) 1 1.667 0.1967
Market 2 5.783 0.0555
Faecal Coliform Treatment (washing) 1 3.636 0.0565

Table 4: Negative log-log regression of Faecal Coliform Presence

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95% Confidence
Faecal Coliform exp(b) Std. Err. z P-value Interval
Market (ref: Madina)
Makola 0.769701 0.350901 -0.57 0.566 0.314968 1.880956
Kaneshie 0.350589 0.147758 -2.49 0.013 0.153480 0.800835

Treatment (ref: Washed)

Unwashed 0.391674 0.166005 -2.21 0.027 0.170670 0.898862

Market#Treatment
(ref:Madina#Unwashed)
Makola#Unwashed 1.615599 0.949898 0.82 0.415 0.510348 5.114476
Kaneshie#Unwashed 3.181185 1.777260 2.07 0.038 1.064243 9.509047

_cons 2.240710 0.755640 2.39 0.017 1.156998 4.339487