Bacterial contamination of babies skin care products in Sudanese markets Yosra.T. Osman* 1, Mohammed Elfatih.A. Omar * 2 1. Department of Microbiology, faculty of pharmacy, Elrazi College for Medical and Technological Sciences, * Corresponding author by email at bitmlookalneel@hotmail.com 2. Al Neelain University, faculty of pharmacy, Dean Office, *Corresponding author by phone 00249912148861. Key words: contamination, skin care ABSTRACT Background: Infants are born with a developing epidermal barrier that is more permeable and more reactive to the environment. Babies skins are very susceptible to infection which may acquire through microbial contaminants in skin care products. Method: Bacteriological quality was examined for 93 babies skin care products, collected randomly from local Sudanese markets. The samples include 39 lotions, 45 talc powders and 9 shampoos, isolation of bacteria passed through many stages, started by inoculation of products into nutrient broth then cultured in selective media and identification of isolates confirmed by using biochemical tests. Results: The incidence of contamination recovered variable bacteria, Bacillus spp isolated from 39 samples (41.9% ), Pseudomonas aeruginosa from 16 (17.2% ), Gram negative rods isolated from 3 samples (3.22%), and a mixture of Gram negative rods and Gram positive rods isolated from 3 samples (3.22%), Staphylococcus aureus was isolated from 4 samples (4.3%), coagulase negative Staphylococci isolated from two samples (2.1%) and one sample was found to be contaminated by E.coli(1.07%). Conclusions: The results showed high incidence of bacterial contamination by potential hazardous microorganisms. Attention must be paid to microbial hazard analysis in critical control points to minimize risk of contamination to produce a product which is microbiologically acceptable. INTRODUCTION The functions of the skin remain essentially the same at all phases of life, including: physical barriers, photoprotection, thermoregulation, immune surveillance, hormonal synthesis, insensible fluid loss prevention and sensory perception (1). The skin of a newborn infant differs from adult skin in several ways that place infants at increased risk of thermal instability, skin damage, percutaneous infection and toxicity from topically applied agents. )2), Immaturity of the epidermal barrier in infant reduces defense against the excessive proliferation of microbes and makes skin more vulnerable to trauma and percutaneous drug toxicity (3). The vulnerability of preterm infant s skin to trauma as there is limited attachment of dermal to epidermal surface, the injured epidermis is a portal of entry of infectious agents, these issues cause difficulties with fluid homeostasis, thermoregulation, toxicity, and infections, As a result, there is greatly increased mortality in premature infants with impaired of skin s functions, generally due to microbial invasion (4); (5). Skin and mucous membranes are normally protected from microbial attack, however protective integuments may be damaged and slight trauma may be result, these situations 36
may be of particular concern when contaminated cosmetics are used in the mucocutaneous membranes or on damaged skin and when used by children under three years, elderly individuals and people showing compromised immune responses.(6) In infants during the first few days of life, the application of certain Gram-negative bacilli to the skin results in colonization and a proportion of the colonized bacteria developed serious infections and fatal generalized infections may include meningitis. The organisms that most commonly responsible of infections in the skin are Staphylococcus spp, Streptococcus spp, and Pseudomonads spp. (6). The overall level of contamination should not only be minimal, but Gram negative bacteria should also be absent in the finished product specially cosmetics use in the skin caring of babies and those used in and around the eyes. (7). Factors such as type of organisms and their virulence, the inoculum size of contaminants, degree of hydration of skin, beside host defense of consumer and presences of trauma or abrasion on skin determine the form of infections (8).There are few reported cases of children s infections due to using of contaminated babies cosmetics.early in 1946 an outbreak of tetanus killed four babies which referred to using contaminated talc powder by Clostridium tetani. (9) Many Studies performed in African countries showed high rate of contamination of cosmetics by Staphylococcus spp, Pseudomonas spp, Clostridium spp, Candida spp and Enterobacteriaceae besides fungi. The main objective of this study to determine the bacteriological quality of babies skin care products in the local markets by isolation, identification and enumeration of specified aerobic bacteria according British pharmacopeia. (10) METHODS A total of 93 samples of babies skin care were products collected randomly from local Sudanese markets, some of them purchased in packets while others were collected singly. The container s labels were recorded, they should include the name and address of manufacturer, ingredients, type of preservatives used, batch number, expiry dates and methods of using for consumers. Experimental 1 ml or 1 gram obtained from samples were added aseptically to 10 ml of nutrient broth, after twenty four hours of incubation a loopful from each nutrient broth tube showing growth was transferred to nutrient agar and then to selective media and incubated at 37 C for 48 hours. The Gram stain technique was performed for colonies obtained from selective media used as cetrimide agar for identification Pseudomonas aeruginosa. Mannitol Salt agar for Staphylococcus aureus. MacConkey agar for E.coli and XLD media for Salmonella. Nine biochemical tests were used to identify bacteria such as Calatase, coagulase and DNase for identification Gram positive cocci and Indole citrate, oxidase, triple sugars fermentation and MR/VP test were used for Gram negative rods bacteria.(10). Total bacterial counting 1gm or 1ml from skin care products was diluted in 9ml physiological saline. A ten -fold serial dilution was made and 1 ml of the second dilutions was inoculated on Plate of Count agar (PCA) in duplicates using the pour plate method. These plates were incubated at 37 C for 72 37
hrs then colonies counts were counted by colony counter. Results were expressed as colony forming unit per gram or ml (cfu/g or ml). RESULTS Out of 93 of samples studied, 43 samples lack batch numbers, 31 without expiry date and tree of preservatives were not written on the labels of products. Lack of batch/lot number should be viewed seriousness as post-marketing surveillance, hence recall, of the product would be difficult to carry out in case of defects. (11). The isolates recovered from these products were Escherichia coli, Staphylococcus spp, Pseudomonas spp. The Bacillus spp, is the most frequently recovered organisms at the initial examination. The incidences of contamination vary between items. Talc powder was found to be more contaminated than aqueous products, 39 out of 45 samples was found to be contaminated. Bacillus spp isolated from 33 samples (73.3%). Pseudomonas aeruginosa isolated from 2 samples (4.4%).Staphylococcus aureus isolated from 3 samples (6.7%) and coagulase negative Staphylococci isolated from one sample (2.2%). Twenty six lotions out of 39 (66.6%) samples studied were found to be contaminated, Pseudomonas aeruginosa isolated from 12 samples (30.7%). Six samples were found to be contaminated with Bacillus spp. (15.3%). Three (7.6%) samples were contaminated with Gram negative rods, three samples (7.6%) by a mixture of Gram negative rods and Gram positive rods (Bacillus spp.), one sample (2.6%) by Staphylococcus aureus and one sample (2.6%) by coagulase negative Staphylococcus spp. Two samples of shampoos were found to be contaminated by Pseudomonas aeruginosa (22.2%).One sample was found to be contaminated by E.coli out of nine samples tested. (11. 1%) Total bacterial counting Table No (1) the result of aerobic bacterial counting of lotions Number of samples Range of aerobic bacterial count Category 1 7 20-100 x 10 2 CFU/ml Category 2 17 100-200 x 10 2 CFU/ml Category 2 2 200-300 x 10 2 CFU/ml shampoo Bacterial count number Numbers Category 1 100-200 x 10 2 CFU/ml 3 Table No (2) the result of aerobic bacterial counting of talc powders Number of samples Range of aerobic bacterial count Category 1 4 100-200 x 10 2 CFU/gram Category 2 29 100-200 x 10 2 CFU/gram 38
Category 3 3 200-300 x 10 2 CFU/gram Category 4 3 >300x10 2 CFU/gram Fig (1): Contamination of Talc powders by aerobic bacteria 39
Fig (2): Contamination of lotions by aerobic bacteria DISCUSSION There are few publications describing the quality of cosmetic products from developing countries but the available evidence suggests that contaminated products are more prevalent, although microbiological quality has generally been improved in recent years. (13). From all contaminants isolated from babies skin care products the presence of Gram negative bacteria particularly Pseudomonas aeruginosa took most concern, it may cause otitis media, UTI, eye and skin infection, In USA a cross contamination cause outbreak of Pseudomonas aeruginosa infections was referred to transmission of this bacteria to neonates from hand lotions used in hospitals.(12). Staphylococci commonly isolated from lotions, creams, moisturizing skin creams, (11) (13) (14), Staphylococcus aureus and Streptococcus pyogenes are most common causes of skin infections. Skin infections cause Staphylococcus aureus is scalded skin syndrome, impetigo, abscesses, cellulites, carbuncles and furuncles, breakdown of the epidermis serves as the entry point for infectious organisms and may be caused by ulceration, trauma, peripheral vascular disease, or preexisting skin conditions that allow bacteria to gain access to deeper tissues, Staphylococcus aureus is capable to secreting several toxins, one of them is leukocidin, this cytotoxin causes destruction of leukocytes and tissue necrosis by inducing production of the potent chemotactic factors.(15) 40
Talc powders In this study isolation of bacteria from talc powders showed, three samples contain Staphylococcus aureus (6.7%), only one contaminated by coagulase negative Staphylococci (2.2%) and two samples contain Pseudomonas aeruginosa. (4.4%), while the majority of bacteria isolated from 33 samples (73.3%). were belong to the Bacillus spp, The results showed difference in types of isolated bacteria, there is low incidence of contamination by Pseudomonas spp and high by Bacillus spp in dry talc powders. High rate of contamination by Gram positive spore forming rods (Bacillus spp) attributed to ability of these bacteria to tolerate desiccation, unlike Gram negative rods bacteria which cannot stand in desiccation.dry powders being mainly susceptible to contamination by Gram positive rods but occasionally by Gram negative rods.(16); (17). In Nigeria Danshen et al., (2011) studied microbial quality of talc powders and found that thirty (50%) of the samples tested were contaminated with Staphylococcus aureus, twelve (20%) were contaminated with Clostridium tetani and four (7%) were contaminated with Candida albicans. Bacillus spp. was also isolated from four (7%) samples while no Pseudomonas aeruginosa was isolated from tested samples. (18). Comparing this study with our results, both studies showed low or absent isolation of Pseudomonas spp. while a high rate of Staphyloccous has been isolated from their samples comparing with Bacillus, and the vice versa in our results. Lotions In South Africa a study was conducted to determine the predominant microorganisms present in spoiled cosmetic creams. The products evaluated included facial creams and body lotions. Pseudomonas aeruginosa and Enterobacter gergoviae were the most predominant bacteria. Of all the samples examined, approximately two third (69%) were microbiologically spoiled. (19). these results agreed with the results obtained from the present work. Twelve samples of lotions out of 39 (30.7%) have been examined and were found to be contaminated by Pseudomonas aeruginosa, that is referred to the high content of water in these preparations which allow bacteria to grow and multiply, Pseudomonas aeruginosa generally prefer high water content so usually aqueous products encourage microbial growth.(19). Okeke and Lamikanra (2001) referred microbial contamination of creams and lotions by Bacillus spp. and Pseudomonas spp. to contamination of raw materials and water as well as the conditions prevalent in the environment where the products were manufactured. Presence of E.coli and other coliforms was due to poor sanitation and high temperatures which increase chance of coliforms to be found in water employed in the manufacture process. (15) Isolation of Staphylococcus aureus from the creams and lotions examined is a function of personnel hygiene, since human skin is a natural source of this organism (12). 41
In Nigeria Okeke and lamikarna (2001), Hugbo et al (2003) and (Osungunna, 2010). Conducted similar studies and all of them showed high incidence of contamination rate by Staphylococcus spp. The results in this study unlike results obtained from the three studies mentioned above; there is low incidence of contamination by Staphylococcus aureus and high by Bacillus spp.(14);(13);(11). Shampoos In the present study 9 shampoos were examined.two of them (22.2%) were found to be contaminated by Pseudomonas aeurginosa and one sample (11.1%) was found to be contaminated by E.coli, while 6 samples (66.6%) were not contaminated. Abdelaziz, A (1989) examined bacterial contamination of shampoos.the results showed predominant contamination by Staphylococcus spp and Pseudomonas aeruginosa while no coliforms were recovered from shampoos. (20) Conclusions Cosmetics always are vulnerable to microbial contamination that attributed to the presence of water and raw materials which usually originate from plants or animals. The findings of this study conclude that babies cosmetics may contain harmful bacteria Presence of organisms such Pseudomonas aeruginosa, Staphylococcus aureus and E.coli in tested products are calling for attention for microbial contents. Microbiological quality of skin products must be controlled in each of the steps and events from the selection of raw materials to consumer use through product formulation, Thus Good Manufacturing Practice (GMP) should be followed, raw materials and water should be tested microbiologically before use, beside using effective preservatives and detect packing and storage ways. It is necessary to carry out routine microbiological control tests for cosmetics after manufacturing and post marketing in order to ensure the quality and safety for customer. Acknowledgements: I would like to express my appreciation and gratefulness to my supervisor Prof/Asim Farouk Mustafa who Guide me patiently throughout the first steps of this research; I ask Alla for him mercy and forgiveness. My deep gratitude to Prof /Mohmmed Elfatih Ahmed Omer for his valuable assistance and generous help. REFERENCES 1. Fluhr, J., Darlenski, R., Taieb, A., Functional skin adaptation in infancy-almost complete but not fully competent, Expermental Dermatology. (2010), 19, 483-492. 2. Siegfried, E., Neonatal sermatology. Rudolph s Pediatrics, New York. The McGraw- Hill Companies, (2002); 21st edition.1168 75. 3. Fernandes, J., Machado. M., Oliveira, Z., Children and newborn skin care and prevention, Anais brasileiros de dermatologia.( 2011), 86,102-10. 42
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