Influence of age at slaughter and sex on carcass characteristics and technological and sensory quality of Goliath chicken meat.

Background and Aim: The Goliath chicken is a slow-growing chicken with a high slaughter weight but whose carcass characteristics and meat quality have not yet been documented. This study aimed to evaluate the effect of age at slaughter and sex on the carcass characteristics and technological and sensory meat quality of Goliath chickens raised in southern Benin. Materials and Methods: Data on the carcass characteristics and technological and sensory quality of meat were collected from 80 chickens raised in confinement and divided into two groups. The first group was reared for 12 weeks of age and the second group was for 20 weeks of age. The animals were individually weighed using an electronic balance (Terraillon 5000g) and then slaughtered. The different parts of the chicken carcass was weighed using the same method. Results: The live, hot carcass, cold carcass, wing, thigh, and tail weights of males were significantly greater than those of females (p < 0.01). The wishbone muscle and thigh and tail assembly pH was lower in the 20-week-old chickens than in the 12-week-old chickens. The brightness and red index of the wishbone of the 20-week-old chickens were significantly higher than those of the 12-week-old chickens (p < 0.001). The red index of the wishbone and the yellow index of the thigh and tail assembly of females were lower than those of males. Conclusion: The meat of 20-week-old Goliath chickens was juicier than that of 12-week-old chickens. Thus, the carcass composition of Goliath chickens is better at 20 weeks of age, especially in males.

On-site verification of the CBEU method using UriSelect4 chromogenic medium at the National Institute of Hygiene (INH) of Lomé.

PURPOSE: The Cytobacteriological Examination of Urine (CBEU) is one of the most requested microbiological analyses in the medical bacteriology laboratory. According to the ISO 15189 standard, Medical Biology Laboratory (MBL) must validate the techniques before their use. It is in this perspective that, within the Medical Bacteriology Laboratory of the INH of Lome, we have carried out the on-site verification of the CBEU method using UriSelect4 chromogenic medium at the INH of Lomé. MATERIAL AND METHODS: The biological material was composed of reference bacterial strains and clinical bacterial strains isolated at INH. Failure Modes Effects and Criticality Analysis (FMECA) with 5 â€‹M model was used to perform risk analysis and performances as repeatability, intermediate fidelity/reproducibility, sensitivity and inter-operator variability were evaluated using references strains and different operators. "Comité Français d'Accréditation" (COFRAC) SH GTA 04 was used as reference. Single-factor ANOVA was used to analyze the data. RESULTS: Operating procedures, Patients preparation, Samples and Preparation of culture media were identified as main critical points with a criticality index of 8, 9, 9 and 12 respectively. The use of uriselect4 to perform ECBU was shown to be repeatable and reproducible. The sensibility of Uriselect4 to detect urinary infection was 100% with negligible inter-operator variability. CONCLUSION: This study performed on-site verification of Uriselect4 at the bacteriology lab of INH of Lome and identified some critical points to master. The overall performance criteria from COFRAC SH GTA 04 were conform.

Description of Antimicrobial Resistance Patterns at the National Institute of Hygiene in Lome, Togo.

The monitoring of antimicrobial resistance has become a priority, mainly in developing countries, to control the emergence of multidrug-resistant bacterial strains. This study evaluated the antibiotic resistance profiles of bacteria isolated at the National Institute of Hygiene in Togo. The laboratory records from January 2013 to December 2015 were examined, which showed that a total of 33,147 samples had been analyzed. Among them, vaginal swabs and urine samples were predominant, amounting to 38.17% and 33.24%, respectively. In total, 3,497 Gram-positive and 1,970 Gram-negative bacterial strains were cultured. Enterobacteriaceae (57.6%), Staphylococcus spp. (21.1%), and Streptococcus spp. (10.5%) were primarily isolated. The records showed that over 3 years, Escherichia coli was resistant to piperacillin (79.66%, 75.1%, and 83%), trimethoprim/sulfamethoxazole (79.3%, 82%, and 82.8%), ticarcillin (89%, 89.7%, and 93%), and amoxicillin (94.1%, 94%, and 96.09%), whereas Staphylococcus aureus was resistant to penicillin G. Streptococcaceae isolates were resistant to trimethoprim/sulfamethoxazole (78.11-87.1%), tetracycline (82.2-91.16%), and norfloxacin (86.16-94.3%). Escherichia coli and S. aureus isolated from urine were more resistant to antibiotics than those isolated from vaginal swabs. There is a need to develop new strategies to fight the emergence of multi-resistant bacteria in Togo.

Hookworm infection associates with a vaginal Type 1/Type 2 immune signature and increased HPV load.

Helminth infection-driven changes to immunity in the female reproductive tract (FRT) is an immune axis that is currently understudied but can have major implications for the control of FRT infections. Here we address how human hookworm infection associates with vaginal immune profile and risk of Human papillomavirus (HPV) infection. Stool, blood, cervical swabs and vaginal flushes were collected from women from the Central region of Togo to screen for hookworms (Ancylostoma duodenale) and high carcinogenic risk HPV types, via Kato Katz and PCR, respectively. Cytokine, chemokine and immunoglobulin levels were analysed in cervicovaginal lavages and plasma samples. A pronounced mixed Type 1/Type 2 immune response was detected in the vaginal fluids of women with hookworm infection and this immune signature was a notable feature in hookworm-HPV co-infected women. Moreover, hookworm infection is positively associated with increased risk and load of HPV infection. These findings highlight helminth infection as a significant risk factor for acquiring a sexually transmitted viral infection and potentially raising the risk of subsequent pathology.

Anti-Th17 and anti-Th2 responses effects of hydro-ethanolic extracts of Aframomum melegueta, Khaya senegalensis and Xylopia aethiopica in hyperreactive onchocerciasis individuals' peripheral blood mononuclear cells.

Hyperreactive onchocerciasis (HO) is characterized by a severe skin inflammation with elevated Th17-Th2 combined responses. We previously demonstrated the anthelminthic activity of Aframomum melegueta (AM), Xylopia aethiopica (XA) and Khaya senegalensis (KS) used by traditional healers to treat helminthiasis in the endemic area of Togo. However, their effect on severe onchocerciasis is poorly investigated. The present study aimed to investigate the anti-Th17 and anti-Th2 effects of hydro-ethanolic extracts of AM, XA and KS during HO. Onchocerca volvulus-infected individuals were recruited in the Central region of Togo in 2018. Isolated peripheral blood mononuclear cells (PBMCs) from both generalized onchocerciasis (GEO) and HO forms were activated with anti-CD3 and anti-CD28 monoclonal antibodies in the presence or absence of the hydro-ethanolic extracts of AM, XA and KS as well as their delipidated, deproteinized and deglycosylated fractions. After 72 hours, cytokines were assayed from cell culture supernatants. Then, flow cytometry was used to investigate the effects of the extracts on cell activation, proliferation, intracellular cytokines and T cells transcription factors. The production of both Th17 and Th2 cytokines IL-17A and IL-5 were significantly inhibited upon T-cell receptor (TCR) activation in the presence of the hydro-ethanolic extracts of AM, XA and KS in HO individuals' PBMCs in vitro. AM and XA inhibited CD4+RORC2+IL-17A+ and CD4+GATA3+IL-4+ cell populations induction. This inhibition was not Th1 nor Treg-dependent since both IFN-γ and IL-10 were also inhibited by the extracts. AM and XA did not interfere with T cell activation and proliferation for their inhibitory pathways. Lipid and protein compounds from AM and XA were associated with the inhibition of IL-17A. This study showed that in addition to their anthelminthic effects, hydro-ethanolic extracts of Aframomum melegueta, Xylopia aethiopica and Khaya senegalensis could downregulate both Th17 and Th2 responses and prevent the severe skin disorder observed.

Hookworm Infections and Sociodemographic Factors Associated With Female Reproductive Tract Infections in Rural Areas of the Central Region of Togo.

Female reproductive tract infections (FRTIs) have a huge impact on women's health including their reproductive health in rural areas. Immunomodulation by helminth infections could influence the occurrence of FRTIs. This study aimed to investigate the association between FRTIs, hookworm infections, and sociodemographic factors in six rural areas of the central region of Togo. A semi-structured questionnaire was used to collect sociodemographical information, and parasitological assessments were used to diagnose helminth infections. Moreover, cytobacteriological examination of vaginal swabs was performed for the diagnosis of candidiasis and bacterial vaginosis (BV), and real-time PCR method was used to determine sexually transmitted infections (STIs). Finally, a logistic regression analysis was performed to assess the relationship and association of these factors to FRTIs. The prevalence of FRTIs was 82.3% including STIs (74.38%), BV (31.79%), and vulvovaginal candidiasis (9.85%). In detail, FRTIs were caused by bacteria such as Ureaplasma parvum (50%), Ureaplasma urealyticum (26.5%), and Mycoplasma hominis (17.5%) and viruses such us cytomegalovirus (5%) and human papilloma virus (HPV) (20%). No cases of Haemophilus ducreyi, Treponema pallidum, or varicella-zoster virus (VZV) were observed. Interestingly, women who had hookworm infections were at high risk of HPV. The use of condoms was a protective factor [adjusted odds ratio (aOR) = 0.23; 95% CI [0.11-0.51)], while the use of contraceptive methods was a risk factor [aOR = 2.49; 95% CI (1.19-5.19)] for STIs. The risk of BV was lower among participants who had more than four pregnancies [aOR = 0.27; 95% CI (0.11-0.65)]. Furthermore, women who had ever been paid for sexual intercourse were at high probability risk of vulvovaginal candidiasis [aOR = 16.92; 95% CI (1.46-196.48)]. This study highlighted risk factors associated with FRTIs, the control of which would help to reduce the incidence of these diseases. Health-care professionals could develop education and sensitization strategies based on these risk factors, and anti-hookworm treatment concepts may be taken into consideration to minimize the risk of HPV infections.

Systemic T Cell Subsets and Cytokines in Patients With Homozygous Sickle Cell Disease and Asymptomatic Urinary Tract Infections in Togo.

Background: In sickle cell disease (SCD), cytokine expression influences the pivotal pathways that contribute to disease pathogenesis. Additional infection could affect the immune profile of patients with SCD and increase disease mortality. The aim of this study was to investigate the cytokines and T helper cells profile in patients with asymptomatic urinary tract infection and homozygous SCD (HbSS). Methods: From July to September 2018, 22 HbSS subjects were recruited at Centre Hospitalier Universitaire Campus in Lomé, Togo, 12 of whom had urinary tract bacterial infections and 10 of whom were uninfected. Cytokines from plasma were measured by the enzyme-linked immunosorbent assay (ELISA) sandwich method, and immune cell profiles were performed by flow cytometry. The immunogenicity of bacteria-derived antigens isolated from the urine of HbSS subjects with asymptomatic urinary tract infections was studied in a cell culture system, and the induction of the cytokines was measured. Results: The mean age of HbSS subjects with urinary tract infections was 20.33 ± 3.58 years, and the male/female ratio was 0.09 (1:11). HbSS subjects with asymptomatic urinary tract infections had elevated plasma levels of interferon gamma (IFN-γ) and interleukin (IL)-10. CD4+Tbet+IFN-γ+ and CD4+FoxP3+IL-10+ T cell populations were decreased in HbSS subjects with asymptomatic urinary tract infections. Bacterial antigens from HbSS subjects induced the production of IL-10 but not IFN-γ in uninfected volunteer donors (HbAA). Conclusion: Our study demonstrated that patients with SCD and asymptomatic urinary tract infections had elevated IFN-γ and IL-10 levels. This chronic inflammatory condition could be a risk for this group of patients in terms of vaso-occlusive crisis. Systematic cytobacteriologic examination of the urine of HbSS subjects would be of interest.

Ethnobotanical survey, anthelmintic effects and cytotoxicity of plants used for treatment of helminthiasis in the Central and Kara regions of Togo.

BACKGROUND: Traditional medicines are the main source of treatment of helminthiasis in endemic areas of Togo. The present study aimed to investigate the plants used by Traditional healers (THs) to treat helminth infections in endemic communities within the Central and Kara regions of Togo and to evaluate the anthelmintic activity of the three most cited plants. METHODS: An ethnobotanical survey was conducted from 19 to 24 June 2017 among traditional healers in the Central and Kara regions of Togo. The anthelmintic activity of the most cited plants namely Aframomum melegueta K. Schum, Khaya senegalensis A. Juss and Xylopia aethiopica A. Rich, was evaluated using microfilariae (Mf) of Litomosoides sigmodontis. The plants were evaluated for cytotoxicity according to the recommendation of NF EN ISO 10993-5 standard using the propidium iodide (PI) dye by flow cytometry on human peripheral blood mononuclear cells. RESULTS: A total of 197 THs were interviewed and 41 plant species were recorded. Leguminosae (14.6%) and Annonaceae (9.7%) families constitute the highest number of species cited for treatment of helminth infections. Afromomum melegueta was the most cited by the THs for the treatment of onchocerciasis (UV = 0.036) while X. aethiopica was associated with the treatment of schistosomiasis (UV = 0.061) and lymphatic filariasis (UV = 0.061). There was a great agreement among the THs regarding ethnomedicinal uses of plants to treat helminthiasis with ICF values ranging from 0.57 to 0.67. The anthelmintic assay yielded lethal doses values of 233 µg/mL, 265 µg/mL and 550 µg/mL, respectively for X. aethiopica, A. melegueta and K. senegalensis. Afromomum melegueta and X. aethiopica presented no cytotoxicity, less than 20% death, whereas K. senegalensis induced moderate toxicity, 24 ± 8% death. CONCLUSION: This study demonstrated the scientific rationale for the use of plants to treat helminthiasis in the Togolese traditional medicine. However, the use of K. senegalensis requires more caution since the plant is fairly toxic. TRIAL REGISTRATION: NA.

ETHNOBOTANICAL STUDY OF PLANTS USED TO TREAT ASTHMA IN THE MARITIME REGION IN TOGO.

BACKGROUND: Asthma is one of the most common chronic diseases in modern society and it is evident that its incidence and severity are increasing, however very little is known about the plants used in the management of the disease. This study therefore aimed to document the plants usage in the Togolese traditional medicine to treat asthma. METHODOLOGY: From January to June 2015, an ethnobotanical survey was conducted using a semi-structured questionnaire with traditional healers (THs) in the southern region of Togo. The importance of the plants species was assessed by the calculated use values. RESULTS: In Total, 121 THs (92 males and 29 females) were interviewed and 98 plants species belonging to 54 families were identified as curing asthma. The most represented families were: Leguminosae with 7 species followed by Euphorbiaceae and Rutaceae contributing with 6 and 5 species respectively. Based on the calculated use values the most important species were Carcica papaya L., Cataranthus roseus L., Eucalyptus camaldulensis Dehnh., Piper guineense Thonn., Eucalyptus citriodora Hook., Eucalyptus globules Labill. and Euphorbia hirta L. The leaves and the root were the parts predominantly used to prepare the formulations, mainly decoctions, administrated by oral route. Clinical manifestations such as wheezing (91.74%), difficulty as speaking or coughing (73.55%), dyspnea (66.94%), dry cough (52.89%), sweating and increased heart rate (52.07%) were used by TH to diagnose the disease. CONCLUSION: This study showed initial evidence of the use of plant materials by Togolese TH to heal asthma. These results could be a starting point for laboratory screenings.

Maize Fungal Growth Control with Scopoletin of Cassava Roots Produced in Benin.

The chemical contamination of food is among the main public health issues in developing countries. With a view to find new natural bioactive products against fungi responsible for chemical contamination of staple food such as maize, the antifungal activity tests of scopoletin extracted from different components of the cassava root produced in Benin were carried out. The dosage of scopoletin from parts of the root (first skin, second skin, whole root, and flesh) was done by High Performance Liquid Chromatography. The scopoletin extract was used to assess the activity of 12 strains (11 strains of maize and a reference strain). The presence of scopoletin was revealed in all components of the cassava root. Scopoletin extracted from the first skin cassava root was the most active both as inhibition of sporulation (52.29 to 87.91%) and the mycelial growth (36.51-80.41%). Scopoletin extract from the cassava root skins showed significant inhibitory activity on the tested strains with fungicide concentration (MFC) between 0.0125 mg/mL and 0.1 mg/mL. The antifungal scopoletin extracted from the cassava root skins may be well beneficial for the fungal control of the storage of maize.

Antimicrobial, Anti-inflammatory and Antioxidant Activities of Jatropha multifida L. (Euphorbiaceae).

BACKGROUND: Jatropha multifida is used in Togolease folk medicine for the healing of chronic wounds. OBJECTIVE: This study aims to investigate antibacterial, anti-inflammatory and antioxidant activities of the leaves ethanolic extract. MATERIALS AND METHODS: The antimicrobial activity was assayed by National Committee for Clinical Laboratory Standards broth microdilution method on strains of Staphylococcus aureus and Pseudomoas aeruginosa isolated from wounds, whereas the anti-inflammatory activity was performed by carrageenan and histamine induced paw edema method in rat modele. The 2, 2-diphenyl-1picrylhydrazyl (DPPH) free radical scavenging and ferric reducing antioxidant power (FRAP) were used for the antioxidant activity. RESULTS: The antibacterial assay showed an in vitro growth inhibition of P. aeruginosa and S. aureus in dose-dependent manner, with minimum inhibitory concentration values ranging from 2.5 to 3.12 mg/mL for S. aureus and from 6.25 to 12.5 mg/mL for P. aeruginosa. The maximum paw anti-inflammatory effect occurred after 3 and 5 h administration of histamine and carrageenan, respectively. The DPPH radical scavenging and the FRAP assays yielded weak antioxidant activity. CONCLUSION: J. multifida possesses antibacterial and anti-inflammatory activities that could justify the use of the plant for the treatment of wounds in the folk medicine. SUMMARY: Antibacterial on germs isolated from wound, anti-inflammatory and antioxidant activities of Jatropha multifida were assayed by NCCLS broth method, carrageenan and histamine, DPPH and FRAP respectively. The results indicated that Jatropha multifida possesses antibacterial and anti-inflammatory and weak antioxidant activities that could justify its use for the treatment of wounds in the folk medicine.

Effects of hydroethanolic extracts of Balanites aegyptiaca (L.) Delile (Balanitaceae) on some resistant pathogens bacteria isolated from wounds.

BACKGROUND: The bark of Balanites aegyptiaca (L.) Delile (Balanitaceae) is widely used in the Togolese folk medicine for the treatment of wounds, malaria and skin diseases. This study aimed to evaluate the antioxidant properties and the activity of the bark extract of this plant against some resistant bacteria isolated from wounds. MATERIALS AND METHODS: The antimicrobial activity was assayed using the NCCLS microdilution while the DPPH free radical scavenging and ferric reducing antioxidant power (FRAP) were used for the antioxidant activity. The susceptibility to conventional antibiotics of bacteria was assessed by an agar disk diffusion method using commercial disk. Phenolics were quantified using spectrophotometric methods. RESULTS: The hydroethanolic extracts of the bark inhibited in vitro the growth of Pseudomonas aeruginosa and Staphylococcus aureus in a dose-dependent manner. Bacteriostatic effects were observed on 15 (33%) and 21 (47%) strains of Staphylococcus aureus and Pseudomonas aeruginosa respectively. The MICs and MBCs against Staphylococcus aureus strains ranged respectively from 2.5 to 12.5 µg/mL and 2.5 to 50 µg/mL. Concerning Pseudomonas aeruginosa, these values were equal (12.5 µg/mL). Multidrug resistance was 95% against six antibiotics for Pseudomonas aeruginosa and 91% against five antibiotics for Staphylococcus aureus. The DPPH radical scavenging and the FRAP assays yielded weak antioxidant activity, closely linked with the total phenolic contents. CONCLUSION: This study indicated that the bark extract of Balanites aegyptiaca possess bioactive compounds implicated in the free radical scavenging and antibacterial activities, justifying the use of the plant in the traditional medicines.

Assessing market-sold remedies in lomé (togo) for hygienic quality.

Traditional concoctions sold in marketplaces are always assumed to be safe and efficient; however, they can be potentially toxic because of poor hygienic practices in plant processing or storage. The present study aimed to assess for the microbial quality of market-sold vegetable drugs in Lomé. Thus, a total of 209 plant remedies were collected in marketplaces and analysed for the presence of total aerobic bacteria, total coliforms, thermotolerant coliforms, Staphylococcus aureus, sulphite reducing bacteria, and yeast and moulds according to the French Association of Normalisation (AFNOR) guidelines. The results revealed that all formulations were contaminated by several microorganisms, excepted alcohol-based mixtures. According to AFNOR limits nonconform drugs were according to total aerobic bacteria (86.96% powders, 81.82% capsules, 66.67% tisanes, and 42.11% decoctions); to total coliforms (9.10% capsules, 8.70% powders and 1.75% decoction); to yeasts and moulds (77.78% ointments, 40% calcined powders, 36.36% capsules, and 23.91% powders). The microbiological analysis revealed that the majority of contaminating bacteria were gram positive catalase positive and oxidase positive bacilli. Quality control studies on market-sold remedies are currently needed to evaluate the microbial risk in consuming these products and they may allow the standardisation of plant processing and storage.

Study of pineapple peelings processing into vinegar by biotechnology.

This study aimed to reduce post-harvest losses of pineapple local variety egbenana by the transformation of juice into vinegar through biotechnological process. Vinegar was produced through two successive fermentations: alcoholic and acetic fermentations. The alcohol fermentation was carried out at 30 degrees C using yeast. Biomass, pH and Brix were evaluated daily during the fermentation. Acetic fermentation was carried out at 30 degrees C using an acetic bacteria strain isolated from pineapple wine previously exposed to ambient temperature (28 degrees C) for 5 days. Biomass, pH and acid levels were monitored each 2 days. The performance of acetic bacteria isolated was also assessed by studying their glucose and ethanol tolerance. The study allowed the isolation of yeast coded Saccharomyces cerevisiae (LAS01) and an acetic bacteria coded Acetobacter sp. (ASV03) both occurring in the pineapple juice. The monitoring of successive fermentations indicated that the pineapple juice with sugar concentration of 20 Brix, seeded with 10(6) cells of Saccharomyces cerevisiae (LAS01) for alcoholic fermentation for 4 days and afterwards seeded with 10(6) cells of Acetobacter sp. resulted in 4.5 acetic degree vinegar at Brix 5.3% and pH 2.8 for 23 to 25 days. The study of glucose tolerance of the strain of Acetobacter sp. showed that the growth of acetic bacteria was important in a juice with high concentration of sugar. However, the concentration of ethanol did not effect on the acetic bacteria growth. These results enabled on one hand to improve the manufacturing technology of vinegar from fruits and on the other hand to produce a starter of yeast and acetic bacteria strains for this production.

Antibiotic resistance in urinary tract bacteria in Ouagadougou.

The present study aimed to ascertain for the current situation of antimicrobial resistance of major urinary tract bacteria in Saint Camille Medical Centre. During two consecutive years, 794 urine specimens were analyzed for microorganism isolation and identification. The microorganisms were identified by conventional methods used in the centre and antimicrobial assays were performed by the NCCLS agar disk diffusion. Pathogenic microorganism's isolation was attempted for 89.04% samples. Escherichia coli (32.76%) was the most frequently isolated microorganism followed by Staphylococcus aureus (22.74%) and Klebsiella pneumoniae (10.45%). The antimicrobial screenings revealed very high antimicrobial resistance, to beta-lactams. The resistance rates recorded with E. coli were 76.64, 74.01, 25 and 74.34% for ampicillin, amoxicillin amoxicillin/clavulanic acid and trimethoprime-sulfamethoxazole, respectively. Microorganisms were still susceptible to quinolones however, attention should be paid, because, the resistance rate already reached 10% for nalidixic acid and ciprofloxacin. Periodic performance of prevalence studies is a useful tool to know the current situation of microorganisms and their resistance patterns in an institution and it helps to access the emergence and the spread of antibiotic resistance.

Hygienic quality of traditional processing and stability of tomato (Lycopersicon esculentum) puree in Togo.

Microbiological and physicochemical qualities of a tomato (Lycopersicon esculentum) puree production line (ripe tomato, washing, cutting, pounding, bleaching, straining, bottling and pasteurization) and its preservation in Togo, West Africa, were studied using the HACCP method. Samples generated during the steps described previously were analyzed by determining sensory, chemical and microbiological characteristics. Samples were analyzed using MPN for coliform populations and plate count methodology for other bacteria. The microorganisms involved in spoilage of the opened products were moulds of genera Penicillium, Aspergillus, Fusarium, Geotrichum, Mucor and gram-positive Bacillus bacteria. The preserved tomato puree exhibited a pH value of 4.3, 90% water content, 0.98 water activity (aw) and an average ascorbic acid level of 27.3mg/100g. Results showed that the critical control point (CCP) of this tomato puree processing line is the pasteurization stage. The analysis of selected microbiological and physicochemical parameters during the preservation of bottled tomato puree indicated that this product was stable over 22 months at 29 degrees C. But the stability of the opened product stored at 29 degrees C did not exceed two months.