Bacillus spp. Contamination: A Novel Risk Originated From Animal Feed to Human Food Chains in South-Eastern Bangladesh

The current study provides information on Bacillus spp. contamination along with present status in commercially available poultry and animal feeds as well as animal-derived products in Bangladesh. The research has been conducted to determine if animal feed and its components are a source of Bacillus spp. contamination in feed and food chain. Out of 180 different feeds, milk, egg, and human stool samples, 218 Bacillus spp. were isolated and identified by cultural morphology, microscopic, biochemical, and molecular characteristics where B. cereus, B. subtilis, B. amyloliquefaciens, B. licheniformis, B. thuringiensis, B. megaterium, and B. coagulans accounted for 51, 22, 9.1, 5.9, 5, 3.6, and 2.2%, respectively. Regarding the enumeration of total viable count and total Bacillus count, correspondingly 67 and 39% samples were found to be contaminated with above 10,000 CFU/g, while highest contamination was 85 and 75% in broiler feed, respectively. The total number of bacteria above the regulatory limits in commercially available feeds indicates a poor compliance with regulation and abuse administration in the Bangladeshi market. Moreover, a hospital-based survey showed that food-borne Bacillus spp. contributed to 4.5% human diarrhea cases and 25% food contamination associated with vegetables, rice, RTE food, milk, and egg, accounting for 46, 34, 14, 4, and 2%, respectively. B. cereus was the dominant isolate correspondingly accounting for 56 and 51% egg and milk contamination followed by B. amyloliquefaciens (32%) and B. thuringiensis (12%) in egg and B. subtilis (25%), B. amyloliquefaciens (12%), B. thuringiensis (6.4%), and B. coagulans (3.2%) in milk, respectively. Toxin gene profiling of Bacillus spp. revealed that B. cereus constituted a principal part of virulence, while B. thuringiensis, B. licheniformis, B. megaterium, B. coagulans, and B. subtilis showed genetic diversity and B. amyloliquefaciens had not carried any toxin gene. Detection rate of enterotoxin genes (nheA, nheB, nheC, cytK, hblA, hblC, hblD, and entFM) showed that 55% isolates carried nheABC genes, 80% entFM, and 71% cytK, whereas only 33% of the isolates contained hblACD gene clusters. These virulence genes were posing a threat to human health due to spread across the food and feed chain. Finally, our findings support the hypothesis that B. cereus might contribute to clinical diarrhea, gizzard erosion, and lung infection in duck and poultry, and that it contaminates animal-derived foods resulting in toxicity and antibacterial resistance to humans. Therefore, maximal tolerance limits of Bacillus spp. and their potential risks to the animal industry are urgently needed to clarify. Moreover, Bacillus spp.–induced toxin residual must be altered for human health via food chain transmission.

Raw Milk as a Potential Source of Food Poisoning Outbreaks

In Africa, the use of poor and unhygienic methods for animal milking and milk processing that leads to spoilage of milk by microbes affects the production of milk and dairy products, especially in the small scale and local processing plants. This study was conducted to analyze the quality and safety of raw milk collected from six different towns in Kwami local government area of Gombe State, Nigeria. The samples were serially diluted using ten-fold dilution and used aliquot 1 ml to inoculate the appropriate media using pour plate technique. The total viable count for bacteria in CFU/ml on plate count agar (PCA) was highest in sample E from U/Anchau with an average of total viable count of 3.8x104 CFU/ml, followed by 3.0x104 CFU/ml in sample B (Dirri), then 2.8x104 CFU/ml in sample D from Burakosuma, 2.5x104 CFU/ml in sample F from Dun urji, 2.3x104 CFU/ml in sample C from Zanbe with least count from sample A at Bele as 1.8x104 CFU/ml. Five (5) bacterial species of public health importance were isolated and identified using biochemical tests namely; Enterobacter sp., Yersinia enterocolitica, Escherichia coli, Staphylococcus aureus and Salmonella sp.. Out of the organisms isolated, Enterobacter sp. had the highest occurrence of 93% (n=280), then Yersinia enterocolitica 90% (n=270), E. coli 70% (n=210), S. aureus 57% (n=170), and finally Salmonella sp. 23% (n=70). Based on the microbiological outcomes, preventive measures for milking and processing that focus on training of farmers and dairy employees for the improvement of the hygiene of local milk and dairy production chain should be defined.

Aspergillus flavus: A degrader of commercial brake fluids

The role of; Aspergillus flavus during the deterioration of brake fluids; was identified. The impact of; A. flavus on used and unused brake fluids; were studied. This research was carried out at the University of Port Harcourt from September 2019 to November 2020. Abro and Jenox DOT3 brake fluids were the samples used for this research. The sample included their unused and used counterpart. Tests to identify microorganisms and physico-chemical parameters; were done. Total viable count, pH and optical density; were tested to check for the level of biodeterioration. Biodeterioration rate was more in Jenox brake fluid than Abro brake fluid. Aspergillus flavus is a major biodegrader in the brake fluid samples. Total viable count and optical density increased as the days progressed during biodeterioration while pH decreased. Brake fluid is an essential fluid needed by most vehicles to function. It is necessary to top-up brake fluid or substitutes; at the appropriate time. No two brands of brake fluid are the same. Therefore, it is advisable not to mix.

Physicochemical and bacteriological screening of pathogenic microorganisms from aquarium water collected from katabon area in Dhaka city

Globally, the ornamental fish business has a high demand for its popularity as well as for its profitability. However, microbial contamination, nutrient depletion and accumulation of organic materials are the major factors in an aquafarming system which can cause death of aquatic animal and causes financial losses. This study aimed to determine the physicochemical and microbiological quality of fish aquarium water. Water of different time interval after treatment (7 days, 20 days, 30 days, 60 days, 90 days) were taken from the different aquariums. A set of physicochemical parameters such as temperature, transparency, pH, conductivity, dissolved oxygen, total dissolved solids, alkalinity, total hardness, conductivity were found within the approved standard concentration for fish. The present study also reveals the microbiological status of the aquaculture water where the total viable count ranging from 2.20×104 to 4.20×104 cfu/ml. Escherichia coli, Staphylococcus spp., and Pseudomonas spp. were present in all samples. Antimicrobial susceptibility tests were also done to determine the antibiotic resistance pattern of microbes isolated from aquarium water. Stamford Journal of Microbiology, Vol.11 (1) 2021: 20-23

The Possible Link Between Manufacturing and Probiotic Efficacy; a Molecular Point of View on Bifidobacterium

Probiotics for food or supplement use have been studied in numerous clinical trials, addressing a broad variety of diseases, and conditions. However, discrepancies were observed in the clinical outcomes stemming from the use of lactobacillaceae and bifidobacteria strains. These differences are often attributed to variations in the clinical trial protocol like trial design, included target population, probiotic dosage, or outcome parameters measured. However, a contribution of the methods used to produce the live bioactive ingredients should not be neglected as a possible additional factor in the observed clinical outcome variations. It is well established that manufacturing conditions play a role in determining the survival and viability of probiotics, but much less is known about their influence on the probiotic molecular composition and functionality. In this review, we briefly summarize the evidence obtained for Lacticaseibacillus rhamnosus GG and Lactiplantibacillus plantarum WCFS1, highlighting that expression and presence of probiotic niche factor (NF) and/or effector molecules (EM) may be altered during production of those two well-characterized lactobacillaceae probiotic strains. Subsequently, we summarize in more depth what is the present state of knowledge about bifidobacterial probiotic NF and EM; how their expression may be modified by manufacturing related environmental factors and how that may affect their biological activity in the host. This review highlights the importance of gathering knowledge on probiotic NF and EM, to validate them as surrogate markers of probiotic functionality. We further propose that monitoring of validated NF and/or EM during production and/or in the final preparation could complement viable count assessments that are currently applied in industry. Overall, we suggest that implementation of molecular level quality controls (i.e., based on validated NF and EM), could provide mode of action based in vitro tests contributing to better control the health-promoting reliability of probiotic products.

Effect of citrus fruit (Citrus sinensis, Citrus limon and Citrus aurantifolia) rind essential oils on preservation of chicken meat artificially infected with bacteria

Essential oils (EOs) obtained from a wide variety of plants have become popular with increased scientific interest as potential natural agents for food preservation. Two concentrations of rind EOs (400 mg/ml and 200 mg/ml) from three species of citrus fruit; Citrus sinensis (Sweet orange), Citrus limon (Lemon), and Citrus aurantifolia (Lime) were used to treat fresh chicken meat inoculated with Escherichia coli ATCC 25922, Salmonella typhi ATCC 20971 and Salmonella enterica ATCC 14028 to evaluate their protective abilities on bacteria-contaminated meat The EOs were extracted from the ground rinds by hydro-distillation. Alongside the EOs, sodium nitrate (NaNO3) was used as a positive control preservative. A viable count was carried out to determine the bacteria load reduction on the inoculated fresh chicken meat. After 24 hours of treatment, the results showed that the EOs had no adverse effect on the physical attributes of the meat: the color and smell of the chicken meat were unaltered compared with the negative control (None EO and NaNO3 treated meat) that showed evidence of putrefaction through color change and foul smell. The two- lime rind EOs concentrations used to treat the Escherichia coli ATCC 25922 inoculated meat reduced the viable count of the organism by 7.9 log compared to the Escherichia coli ATCC 25922-inoculated meat which received no rind EOs or NaNO3 treatment. Other results showed that sweet orange (SO) rind EOs (400 mg/ml and 200 mg/ml) treatment of meat inoculated with Salmonella enterica ATCC 14028 had similar but mild preservative effects as both treatments reduced the log of the bacteria by 1.1 and 0.8, respectively. In comparison with NaNO3, the EOs treatment had a significant (p<0.05) preservative effect on the bacteria-inoculated meats. Findings from this study, therefore, suggest that Citrus spp. rind EOs have good potential as natural preservative for chicken meat. However, notwithstanding the relative positive organoleptic results observed in this study, further investigations on the prolonged preservation effect of the EOs on the physical attributes of fresh chicken meat need to be undertaken.

The pencil eraser swab technique to quantify <i>Cutibacterium acnes</i> on shoulder skin

Introduction: Cutibacterium acnes is the most common cause of postoperative infections in orthopaedic shoulder surgery and is hard to eradicate with current measures. Newer strategies focus on reducing bacterial load on the skin before surgery. Several previous studies have used a large number of both described and undescribed sampling techniques. The purpose of this study was to compare three previously described swab techniques to obtain bacterial cultures: Levine's (L) technique, the Z technique and the pencil eraser swab (PES) technique. Methods: Three consecutive skin swabs were collected from the right shoulder, on 15 healthy male volunteers, using Levine's technique, Z technique and PES technique from each participant. To determine the number of living bacteria, serial dilutions were made, and after culturing for 5 d, viable count (VC) was expressed as CFU/mL (with CFU representing colony-forming unit). Results: The PES technique yielded significantly higher VC than the two others. PES: median 3700 CFU/mL, L: 200 CFU/mL and Z: 220 CFU/mL (p=0.003). There was no significant difference between the methods regarding the number of positive cultures. PES: 14/15, L: 11/15 and Z: 12/15. Conclusions: There is a need to harmonise sampling techniques of C. acnes in order to compare the efficacy of different measures to reduce the bacterial load on the skin before and during surgery. Of the three tested methods, the PES technique is simple and produces the highest bacterial counts.

Molecular identification and technological properties of yeasts isolated from spontaneously fermented cassava waste pulp

The aim of this work was to report on molecular identification and technological properties of the yeast flora isolated from spontaneously fermented cassava waste pulp. This was done with a view of obtaining yeast strains that could be used as a starter culture for the fermentation of cassava waste pulp. Molecular identification was based on the nucleotide sequence of the ITS region of the genomic DNA of the yeast isolates while the technological properties evaluated include linamarase (U/mL), gelatinase, and haemolytic activity; growth at pH 2.5 and tolerance to 2 % bile salt. All the representative five isolated yeasts were identified as Geotrichum silvicola KLP04 – KLP08. The isolates exhibited linamarase activity ranging between 3.3 and 4.2 with strain KLP04 having the highest value and strain KLP05 the least. None of the isolates demonstrated gelatinase and haemolytic activity except strain KLP08 which was partially haemolytic. All the examined yeasts exhibited good growth at pH 2.5, with strain KLP08 having the highest viable counts of 4.1 log10cfu/ml and strain KLP04 the least value of 3.5 log10cfu/ml after 72 h of growth. All the identified yeasts showed strain-specific tolerance to 2% bile salt with strain KLP04 having the highest viable count of 4.3 log10cfu/ml and strain KLP08 the least value of 2.2 log10cfu/ml at the end of 72 h of incubation. Based on all the examined technological properties, Candida silvicola KLP04 strain had the highest potential to be considered for starter culture for the fermentation of cassava waste pulp.

Physicochemical and microbiological evaluation of surface water quality of aquaculture ponds Located in Savar, Dhaka, Bangladesh

Aquaculture is one of the most vital sectors in Bangladesh as it exhibits a major role in nutrition, livelihoods and foreign exchange incomes/earnings every year. However, due to chemical impurities, infectious diseases caused by microorganisms, heavy metal accumulation, and aquaculture in Bangladesh is gradually declining and posing serious health risks. In Savar, which is one of the major industrial zones in Bangladesh, all industrial sewage and wastes severely deteriorate the water quality of the ponds, rivers, lakes and various waterways that are involved in aquaculture/fish culture. Hence, to determine the water quality by assessing different physicochemical and microbiological parameters, water samples were collected from five selected ponds located in Atomic Energy Research Establishment premises, Savar, Dhaka and analyzed according to the standard procedures. The obtained values of temperature, pH, Salinity, TDS, TA, EC, TH, Chloride content, Free CO2, DO, Nitrate and Sulfate were compared with the recommended values of Bangladesh and WHO standard for suitable water quality. Most of the physicochemical parameters exceeded the Standard value. Total Viable Count, Total Coliform and Fecal Coliform Count were also found to be higher than the standard value of WHO indicating fecal contamination of the pond water. Some fish pathogens were also isolated from the ponds. Water quality index (WQI) was calculated for five sampling sites to determine the level of pollution. It was observed that the water quality of the all the ponds reached to critical point of pollution. It is therefore, a high time to take initiatives to save the ponds that are involved in aquaculture from further pollution. The results revealed that the pond waters of five different sites were excessively polluted and unsuitable for fish culture.

Plant Extracts for the Control of Listeria monocytogenes in Meat Products

The antimicrobial activity of garlic (Allium sativum L.) and onion (Allium cepa L.) plant active extracts was determined against Listeria monocytogenes in two meat products. Samples of sausages “cacciatore” and cooked ham in vacuum-packaged slices were artificially contaminated, and the presence of Listeria was evaluated during the sausages ripening and throughout the shelf-life of the cooked ham. The test carried out on sausages did not show differences among treated and untreated samples. The antagonistic activity of the plant extracts against the pathogen was probably hidden by the competition from the sausages microbial flora and the pH and the water activity (aw) decrease. On the other hand, the plant extracts determined an initial reduction of about 1.00 log cfu/g of the L. monocytogenes viable count in the cooked ham slices contaminated with 103 cfu/g, but the best result was obtained with the contamination of 102 cfu/g of L. monocytogenes. In addition to the pathogen’s initial decrease, we observed an extension of the lag phase and a reduction of the Listeria growth rate. Considering that the presence of L. monocytogenes during the slicing phase of the cooked ham does not exceed 10 cfu/g, the use of plant extracts can lead to complete pathogen elimination.