Abstract: Spices have been get Great attention among the world population because of its great value in food floor and aroma. Moreover it is benefiting for sellers by creating job opportunities. However it perceived to be major public health problem due to microbial contamination. The purpose of this study was to evaluate microbial safety of spices in Jimma town. The study involves laboratory analysis for microbiological safety of spice. The standard method were used for the enumeration of Aerobic mesophilic, Entrobacteriaceae, coli forms, Staphylococci, yeasts and molds. A total of 40 spice samples Red chilies (Capsicum spp.), Turmeric (Curcuma longa) were collected from Mercato, Kochi and Agip selling sites. This result show that the overall mean microbial counts (CFU g-1) were dominated by Aerobic mesophilic bacteria (7±o.4), yeasts (6±o.4), coliform (6±0.0), molds (6±0.0), Staphylococci (4±0.8) and entrobacteriaceae (5.5±0.5). Out of total 208 isolates characterized, the most dominant were Bacilli spp. (83.33%) followed by Staphylococci spp. (14.3%). Out of 20 mycopopulation isolates characterized, molds were dominant (75 %) and yeast were 25%. Among the molds isolates, Mucor spp. were predominant (41.6%) followed by Aspergillus spp. (16.6%) whereas Pencillium and Geotrichum spp. (12.5%) while Eurotium, Rhizopus and Fusarium spp. were least recorded fungi (8.33%). Spices contamination problem in the present study could be due to poor preparation, selling site, storage area and materials and poor personal hygiene. Generally, the microbial safety of spices sold in Jimma town was poor and call for special attention.
INTRODUCTION
Spices are non-timber forest product which extract or obtained from plant vigor such as seeds, fruits, flowers, rhizomes, bulbs, bark, leaves and stems that used as food additive in order to provide odor, smell, taste flavor (Leek et al., 2004). It encompasses of different chemical nutrient and minerals including water, protein, fat, carbohydrates, ash, calcium, potassium, sodium, phosphorus, iron and various essential vitamins (Singh et al., 2002). Spices are essential components of cuisines since ancient and are used in minute amount impart aroma in food preparation to improve their palatability (Hampikyan et al., 2009). They are also used for stabilizing several food items from deterioration (Mandeel, 2005). It has also been prescribed for aiding digestion, raising sexual potency, decreasing blood pressure, controlling metabolism and delaying the onset of degenerative disease (Jayaprakasha et al., 2001). On the other hands, Spices used as refreshing food, drink and daily meal components (Anderson et al., 2000). Nowadays, Spices are known to have some ethno-botanical medicine and antimicrobial properties (Singh et al., 2002; Knefiel and Berger, 1994). Plants traditionally used for medicinal purpose in different parts of the world have been screened for possible antimicrobial action by severaworkers (Bonjar, 2004). This value of spices also reported by many scientists (Leite de Souza et al., 2006; Karaman et al., 2011). The combined value of spices also generally show synergistic antimicrobial effect specially on fungus (Das, 2002). This is due to the presence of different phyto-chemicals or compounds like alkaloids and terpenoids (Jackson et al., 1995).
Even if, spices are multipurpose non-timber frost products it could be spoiled and contaminated by pathogenic microbes during and post-harvesting in poor hygiene condition (Gurbuz et al., 2000). Hence, they are considered as significant carriers of microbial contamination primarily molds and some bacteria (Dimic et al., 2000; Romagnoli et al., 2007). Therefore, Spices are a primary alimentary intoxication when added to foods in which the pathogen is favorable to grow (Hampikyan et al., 2009). According to Vij et al. (2006), the possibility of pathogen growth in higher rate in spices in case of poor hygienic condition when it used in food without subsequent preservation and that may lead to serious disease. Similarly, several studies show that spices contain various types of microorganisms such as pathogenic and toxigenic molds (Banerjee and Sarkar, 2003). Moreover, Martins et al. (2009) explained that molds from spices may reduce the quality of food and creates potential risk for human health with the production of toxic metabolites called mycotoxins. To this effect, the present study was designed to assess the microbial safety of some selected spices in Jimma town with the following objectives.
MATERIALS AND METHODS
Study area: This study was conducted in Jimma town located at 353 km south west of Addis Ababa the capital city of Ethiopia. The geographical location of the town is 7°41`N latitude, 36°50` E longitudes and an average altitude of 1780 m above sea level. The average minimum and maximum Temperature of the town is 14 and 30°C, respectively with annual rainfall ranges from 1138-1690 mm (Alemu et al., 2011). This study was covered from September-may 2013.
Sample collection: A Total of 40 samples were collected from spices sellers of Jimma town, from September-May 2013.The sample was purchased from spices sellers at 2-5 AM. The sample was transported to Research and post-graduate laboratory Department of Biology, College of Natural Sciences and Jimma University main campus by investigator. The spices were kept in the Refrigerator at 4°C until analysis was conducted.
Sample preparation and microbial enumeration
Sample preparation: A 25
g of spices sample was suspended in 225 mL of saline solution and homogenized
in erylenmer flask for 5 min using shaker at 160 rpm. A 1 mL of homogenized
sample was transferred in 9 mL of saline solution and mixed thoroughly by using
vortex mixer. The homogenized spices sample was diluted from 10-1-10-8
and 0.1 mL of aliquot dilution was plated on pre-solidified plate and incubated
at appropriate temperature (32°C) for 18-48 h. The colonies were counted
from plate containing microbial colonies between 30-300 and expressed in colony
forming unit per gram (CFU g-1).
Microbiological analysis
Determination microbial spectrum: For Spice sample content Total Aerobic
Mesophilic Counts (TAMC), Counts of Staphylococci, Count of Enterobacteriacea,
Bacillus, Counts of yeast and molds and Counts of Coliforms were determined
according tothe criteria specified in (Dabassa and Bacha,
2012) and by standard plate count methods on PCA (Plat Count Agar) (Oxoid)
for TAMC at 32°C during 72 h, Mannitol Salt agar (Oxoid) for staphylococci
count,Chloramphenicol Bromophenol Blue Agar for yeast and mold and VRBGA (Violet
Red Bile Glucose Agar) (Oxoid) for Enterobacteriacea and VRBA (Violet Red Bile
Agar) (Oxoid) for Total Coliform Count (TCC) during 24 h at 32°C for TCC
and 44°C for FCC.
RESULTS AND DISCUSSION
Microbial counts: Spices may be in corporate to processed food or may be added as an ingredient or condiment to food prepared in home or food businesses. It may have a high incidence of microbial contamination. The International Commission on microbiological specification in foods recommended that spices can be treated as a raw agricultural commodity with ultimate use of the product determining the acceptable microbial load (Omafuvbe and Kolawole, 2004). In addition, spices have implicated in large scale outbreaks of food borne illness (Frisvad et al., 2005). Additionally, utensil incase container of spices used by sellers has also its own impact in facilitating spice contamination and may have originated from sellers (vendor) hand when touch items uncover (dishcloths) (Cardinale et al., 2005). So that, the mean count of Aerobic Mesophilic Bacteria (AMB) was highest (8.23 log CFU g-1) in red chillies and lower (6.6 log CFU g-1) in Turmeric (hird). The heat resistant spores of spore former bacteria may survive cooking while vegetative bacteria were eliminated (Freire and Offord, 2003). Similarly Fufa and Urga (1996) stated the food items that sold at stalls have spore forming bacteria which are the main concern due to temperature time abuse. The mean count of coliforms was high (6.2 log CFU g-1) in Turmeric and low (5.8 log CFU g-1) in Berber whereas mean count of Entrobacteriaceae low in red chillies (5.3 log CFU g-1) while high (5.8 log CFU g-1) in Turmeric. However, the mean count of Staphylococcus aurous was as high as red chillies (4.8 log CFU g-1) in Turmeric and low in red chillies (Berbere). On the other hand, the mean count of yeast was high (6.9 log CFU g-1) in the turmeric whereas low (5.8 log CFU g-1) in the red chillies. But the mean count of molds was high (6.2 log CFU g-1) in red chillies and low (5.8 log CFU g-1) in turmeric. The reason why a wide range of spice contaminates with the moulds is that its spore can be carried out by wind and hence can easily enter into good sample (El-kady et al., 1995).
The highest mean counts of AMB were recorded in spice sample red chillies from Mercato (10.3 log CFU g-1) and turmeric from Kochi (7.75 log CFU g-1). The mean count of total Aerobic mesophilic bacterial observed in the present study is line with Craig (1999) reported the total plate count of AMB in Red chilies’ greater than 1x107 CFU g-1 from Australian spices. Similarly, the highest mean counts of Entrobacteriaceae in spices sample from Mercato (5.5 log CFU g-1) and turmeric from Kochi (5.1 log CFU g-1) and highest mean counts of coliforms were recorded from sample Turmeric of (Mercato)(6.9 log CFU g-1) and Red chillies from Kochi (5.8 log CFU g-1) whereas highest mean of yeast and molds were recorded in spices sample of Turmeric from Kochi (7.88 log CFU g-1) and Red chillies from Kochi (6.8 log CFU g-1) and Turmeric from Kochi (7.47 log CFU g-1) and Red chillies from Kochi (6.5 log CFU g-1), respectively. Furthermore, the highest mean count of Staphylococcus aereus was recorded in samples (turmeric) from Kochi (5.1 log CFU g-1) and red chillies (Berbere) from Kochi area (4.9 logCFU g-1).
The maximum and minimum mean counts of both spices samples were shown in (Fig. 1). Accordingly the highest mean counts (log CFU g-1) of AMB, coliforms, yeast, molds, Staphylococcus aereus and Entrobacteriaceae were recorded in spices samples. The mean count of Entrobacteriaceae in the present study was 5.5 CFU g-1 while the coli form count was 6 CFU g-1 irrespective to (Colak et al., 2006) who reported as 3.79 CFU g-1 from turmeric.
| Fig. 1: | Mean maximum and minimum microbial count of red chilies spice sample collected from different site, Jimma town |
The mean count of AMB was highest (10.3 log CFU g-1) in red chillies sample collected from Mercato site and the mean counts of coliform was 5.8 log CFU g-1. The mean counts of Entrobacteriaceae in the same sample was 5.5 log CFU g-1 followed by Staphylococcus aereus 4.8 log CFU g-1 but the highest mean count of yeast and molds was 4.4 and 4.7 log CFU g-1, respectively. On the other hand the highest mean count of AMB from Mercato turmeric was 6.2 log CFU g-1 but the Entrobacteriaceae mean count was highest (6.5 log CFU g-1) than AMB count. However, the mean count of coliform, yeast, molds and Staphylococcus aereus were 6.0, 5.5, 4.45 and 4.55 log CFU g-1, respectively as in Fig. 1.
The results also show that the maximum mean microbial count (log CFU g-1) of yeast, molds, AMB, Entro, coliform and Staphy in red chillies collected from Kochi site were 9, 8.7, 6.9, 6.2, 6.07 and 5.04, respectively, meaning they were above detectable level as shown in (Fig. 1). But it is line with (Chandarana et al., 2005) who reported the high count of AMB Turmeric to be greater than 2.5x105 CFU g-1 from Japan spices. Similarly, the maximum mean microbial count (log CFU g-1) of AMB, yeast, molds, coliforms, Staphylococcus aereus and Entrobacteriaceae in turmeric collected from Kochi site were 9.5, 9.43, 8.47, 6.64, 5.3 and 6.1, respectively. They were also above detectable level in samples as it expressed in (Fig. 2.) In contrast to this to this, the recent study indicate that the total count of AMB was greater than the limit.
Only the mean count of yeast and molds were detected from Agip site. Accordingly, the highest mean of yeast count in red chillies and turmeric was 6.5 and 7.3 log CFU g-1, respectively. The mean count of yeast and molds in the current study is in line with Nair and Chanda (2006) who reported counts between 2-6 CFU g-1 from prepacked of Indian spices. However, the highest mean count of mold in red chillies and turmeric was 7.3 and 5.4 log CFU g-1, respectively as it shown in (Table 1). In contrast to this study Buckenhuskes and Rendlen (2004) reported that count of yeast and molds to be 3.08 CFU g-1 from species in chain stores. INSC also reported that the maximum count number of molds has to be 5x103 CFU g-1 in contrasts to present study.
| Fig. 2: | Mean maximum and minimum microbial count of turmeric spice sample collected from different site, Jimma town |
| Table 1: | Mean microbial count minimum, maximum (log CFU g-1) |
Mycopopulation analysis and characterization: From total of 20 fungal populations isolated Mucor spp. Were the predominant fungus in both Turmeric (50%) and Red chillies (33.33%) followed by yeast (25%) in red chilies and 25% in turmeric (hird). On the contrary, both Pencillium and Goetrichum spp. were the less dominant fungal strain isolated from turmeric (12.5%) and Aspergillus species were 16.6% while Rhizopus, Eurotium, Fusarium spp. were the least dominant fungal species isolated from red chilies (8.33%) (Table 2). Aspergillus, Rhizopus and Fusarium were the least dominant fungal isolates. In similar to the present study of Avato et al. (2000) who reported Aspergilus spp. isolated from Red chillies.
The Fusarium, Rhizopus and Aspergillus spp. were isolated from both turmeric and Redchillies that a agreed with Shadanaika (2005) from India. Similarly, Joe et al. (2009) reported that Rhizopus spp. was found in Turmeric powder.
Microscopically observed and characterized of these fungi indicated in (Table 3). Generally among the isolates fungal strain Mucor spp. were predominant (41.66%) followed by yeasts (25%) while Aspergillus spp. were (16.6%) and Goetrichum and Pencillium spp. were (12.55%).
| Table 2: | Frequency distribution of dominant fungi (molds and yeasts) from the selected spices, in Jimma town, south western of Ethiopia |
| Table 3: | Mycopopulation distribution of dominant fungi in the selected spices, Jimma town, south western Ethiopia |
Fusarium, Rhizopus and Eurotium spp. were (8.33%).
CONCLUSION
Spices are sourced from the roots, stems and seeds of the aromatic plants and sold at the market place. It can be become contaminated with pathogenic microorganisms during growth, harvesting, post harvest processing and during selling in the market place. The overall microbial quality of species assessed in this study was very poor, as sellers put spices utensil on the floor which easy to soil borne contaminate to it. Furthermore, the most predominant microbial group was bacillus spp. Staphylococci, Penicillium and Aspergilus spp. Thus the presence of this microbial group could be a possible predication for the presence of pathogen. Thus, the presence of these microbial groups could be a possible means to produce toxic substances that cause disease or illness to human being upon consumption.