Prevalence of Moulds in Households Drinking Water of Some Local Government Areas of Kano, Nigeria

ABSTRACT


INTRODUCTION
Water is one of the most abundant and essential commodities of man occupying about 70% of the earth's surface, yet a greater percentage of the world's population, most especially in developing countries live without access to safe water (Hazen and Toranzos, 1990;Adriano and Joana, 2007).Nigeria for example, is located in coastal West Africa where water is abundant, yet most of the population lacks adequate and safe drinking water.This thus prompted the sinking of boreholes by rich individuals and selling the water to the ever growing population without any treatment.Many Nigerians are engaged in packaging water, popularly called "pure water" in polythene bags of about 60 -65cl and selling to the public.The safety of this "pure water" is still questionable because many who are engaged in its production do not follow strictly the standards set by FEPA (1999) and WHO (2006) for safe drinking water.
Fungal infections are becoming more and more important because of increasing numbers of immune suppressed patients.Nonetheless, waterborne fungi are associated with taste and odour problems, contamination of food and beverage preparation, and in a variety of health related effects (Nagy andOlson, 1982, 1985;Hinzelin and Block, 1986;Geldrich, 1996;Doggett, 2000;Joseph and Michelle, 2003).
A wide variety of fungi species have been isolated from water in various investigations.The lists of taxa reported in these investigations vary from study to study.West (1986) demonstrated that fungi isolated from portable water were dematiaceous (63%) and more especially Cladosporium (27%), Phoma (9%), Alternaria and Exophiala (each 7%).Arvanitidou et al. (1999), reported Penicillium, Aspergillus and Candida as the major genera isolated in their study while Ana et al. (2006), indicated Acremonium (38.2%) and Penicillium (40.59%) as the major isolates amongst others in tap water in Braga, Portugal.Gunhild et al. (2006), found Penicillium spp, Absidia spp, Acremonium spp, Aspergill us spp and Mucor spp to be the major fungi genera inhabiting Norwegian drinking water.Some of these species isolated from water samples are known to be strong allergenic skin irritants or may cause infections in immune suppressed individuals such as AIDS, cancer, organ transplant patients and persons with asthma or various respiratory problems (Gunhild et al., 2006).An increase in the number of invasive diseases due to fungi has occurred recently (Arvanitidou et al., 1999;Anaissie et al., 2003).In Nigeria, water borne diseases are one of the main problems in rural and urban communities.These diseases are as a result of bacterial, fungal or other microbial infection of water.Unfortunately , most water screening methods in Nigeria are focused on the occurrence and significance of bacteria with little attention to other microorganisms such as fungi.It is on this note that we decided to investigate the prevalence and significance of moulds in household's drinking water in Kano state, Nigeria.

Study area
The study was conducted in four local government areas of Kano State viz: Ungogo, Dala, Kunbotso, and Gwale.

Sample collection
Standard method described by American Public Health Association (APHA, 1999), was used for the collection of samples.During collection of samples, in each targeted house, 300ml of water was poured aseptically in to 300ml sterilized bottles.For tap water and borehole, the samples were collected by allowing the water to run to waste for 2 or 3 minutes and then the water were aseptically collected in sterile bottles.Water from wells was collected by means of a sterilized bottle fitted with a weight at the base.All samples collected were then labeled with sample number, date of collection and sample source for analysis purposes, and then sealed.After sampling, a structured questionnaire was administered to each participating household.The questionnaire included variables such as family size, devices used to collect and store water, storage duration and water treatment.Samples collected were then transported to the laboratory in an iced cooler for storage as soon as possible.

Sample filtration
Membrane filter assembly was set up by inserting the glass funnel bottom in to the opening of arm jar flask.At the side of the flask, there is narrow opening; this was then connected to the vacuum pump machine through rubber tubing.During filtration, membrane filter was placed in to the funnel using sterile forceps.Sample was shaken vigorously at least 25 times up and down to mix the sample and then 100ml of sample was poured in to the funnel and the vacuum pump was then turned on to drain the sample through the sterile 47mm and 0.45µm membrane filters (Whatman, Maidstone, Japan).After filtering the sample, the funnel walls were rinsed three times with 20-30ml of sterile peptone water, then the vacuum pump was turned off and the funnel top was lifted up to remove the membrane filter using sterile forceps and the filter was placed on Sabouraud Dextrose Agar, followed by incubation at 20-25 0 C for 3-5 days (APHA, 1999).

Isolation and identification of mould
Colonies on SDA were sub cultured in the same medium to isolate a pure single colony for identification test.During identification, a drop of 95% ethanol was placed on a microscope slide.Using a sterile inoculating needle, a small portion of fungal growth was gently removed and placed on 95% ethanol and then gently spread it out with two dissecting needles so that it can easily be identified when viewing.When most of the ethanol has evaporated, a drop of lacto-phenol cotton blue was added and covered with a cover glass and then examined microscopically (Chei et al., 2000;Food and Agriculture Organization, 1979).

DISCUSSION
In the study, the most frequently isolated mould was the genus Aspergillus.These findings are consistent with the works conducted by Arvanitidou et al. (1999 and2000) and Gunhild et al. (2006), that Aspergillus was the most commonly isolated genera in water.Aspergillus are known to produce aflatoxins (B1, B2, G1 and G2), the most toxic and potent hepatocarcinogenic natural compounds ever characterized (Bennett and Klich, 2003).These fungi cause a wide range of diseases in humans, ranging from hypersensitivity reactions to invasive infections associated with angio-invasions.A. niger, A. terreus and A. nidulans where found on several occasions during this study.The finding of A. niger was in agreement with the work conducted by Hageskal et al., (2006) in which he also reported the frequent occurrence of A. niger in drinking water.However, the presence of A.terreus and A. nidulans was not consistent with the work of Okpako et al., (2009) in that he reported the frequent occurrence of A. flavus in borehole water samples.A. niger is a common allergen and may cause opportunistic invasive infections in hospitalized immunized patients (De Hoogetal, 2000).
Penicillium species were especially abundantly distributed and clearly have the ability to survive water treatment and contaminate water reaching various network installations.Only heating of water seems to inhibit the recovery of viable Penicillium spores or hyphae.The implication of Penicillium species in allergy, asthma, or other respiratory problems has been a subject of several studies worldwide (Schwab and Straus, 2004).Strong associations between Penicillium spp.and health problems were also reported by Cooley et al. (1998).Hence, many of the species isolated in the present investigation may have allergic potential if susceptible individuals are exposed.Furthermore, several of the demonstrated Penicillium species have been reported to be active mycotoxin producers (Frisvad et al., 1998, Moreau, 1979, Samson et al., 2004).This fact raises the question of potential mycotoxin production in water and further investigations into this problem are merited.The genus Penicillium also includes common contaminants of food and beverages (Pitt andHocking, 1999, Samson et al., 2004).It is not unlikely that water can be the route of transmission for mold contamination and spoilage of foods.
Fusarium species were also found in large numbers 40(19.4%).Several researches have been found to show the occurrence of Fusarium species in water environments.Okpako et al., (2009), reported the occurrence of Fusarium species on several samples 16(15.7%).Similarly, Anaissie et al., (2002), recorded 14 species of Fusarium from water sampled in hospital.Fusarium species have been recognized as agents of superficial infections (keratitis and cutaneous infections, onychomycosis and infections of wounds and burns) (Guarro and Gene, 1995).In recent years, deepseated and disseminated infections have been increasingly described in immune compromised patients, especially in neutronpenic patients (Guarro and Gene, 1995).The prognosis is very poor and death occurs in up to 70% of cases despite antifungal therapy (Musa et al., 2000).
Very small percentage of Rhizopus was found in the research.Okpako et al., (2009), recovered a significant percentage of Rhizopusin sachet and borehole water.Zygomycetes are known to cause diseases in immune compromised patients (Sheppard et al., 2004 andAna et al., 2006).The genus Mucor is known to be a major cause of thrombosis, infarction, nasal or paranasal sinus infection and GI disorders.
The presence of these filamentous fungi may be mainly associated with post treatment contamination from outside sources, or post collection contamination, or from populations growing within biofilms or other materials (such as pipe joints and seals) in the distribution system, or they were able to escape the treatment procedures or the contamination is the source.(Bays et al., 1970, Grabinska et al., 2007, Ramirez-toro and Minnigh, 2002).
The detection of pathogenic microorganisms in different sources of drinking water also reveals the alarming situation for households' water.The high prevalence of filamentous fungi in households' drinking water is a matter of serious concern.
On comparison, there was no significant difference in the occurrence of moulds between water collected from boreholes, households, taps and wells (p>0.05).Also a significant difference was not found in the occurrence of moulds in waters collected from Kumbotso, Gwale, Dala and Ungogo (p>0.05).The lack of a difference between these sites may be associated to the fact that most of them have the same sources of water.
The mean number of cfu/100ml was 4.50, 3.60, 3.13 and 3.43 for boreholes, households, taps and wells respectively.Yamaguchi et al., (2007), also reported the mean number of cfu/ml as 10.8 and 11.6 for yeasts and moulds in bottled mineral water, respectively.In municipal tap water, he also reported 2.8cfu/ml for yeasts and 1.0cfu/ml for moulds.
It is unlikely that the occurrence of molds in water at the concentrations observed in this study would cause disease in healthy individuals.However, if the right conditions are present and regrowth of molds occurs in water systems, exposure of humans to large amounts of potentially harmful mold species could become a problem.Several of the molds are potential toxin producers and exposure to small amounts of toxins for several years may have negative effects on the immune system (Letscher-Bru, et al., 2002, Vismer, et al., 2002).

CONCLUSION AND RECOMMENDATIONS
Since most of the species of molds isolated are pathogenic to humans, it is likely that exposure of humans to very large amounts of potentially harmful mold species could cause infections to healthy individuals.Owing to the health hazards associated with the fungi isolated from drinking water used for this study, the following recommendations are necessary; Prevention of storm flooding into springs and wells.If source water is microbiological clean, then use of containers with a narrow mouth and lid, would render boiling unnecessary.Governments should improve dissemination of information on private water testing, personal hygiene and sanitation.