Microbial Contamination
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2021 ◽  
Vol 19 (1) ◽  
Victoria Ann Remley ◽  
Jianjian Jin ◽  
Sarmila Sarkar ◽  
Larry Moses ◽  
Michaela Prochazkova ◽  

Abstract Background Gene transfer is an important tool for cellular therapies. Lentiviral vectors are most effectively transferred into lymphocytes or hematopoietic progenitor cells using spinoculation. To enable cGMP (current Good Manufacturing Practice)-compliant cell therapy production, we developed and compared a closed-system spinoculation method that uses cell culture bags, and an automated closed system spinoculation method to decrease technician hands on time and reduce the likelihood for microbial contamination. Methods Sepax spinoculation, bag spinoculation, and static bag transduction without spinoculation were compared for lentiviral gene transfer in lymphocytes collected by apheresis. The lymphocytes were transduced once and cultured for 9 days. The lentiviral vectors tested encoded a CD19/CD22 Bispecific Chimeric Antigen Receptor (CAR), a FGFR4-CAR, or a CD22-CAR. Sepax spinoculation times were evaluated by testing against bag spinoculation and static transduction to optimize the Sepax spin time. The Sepax spinoculation was then used to test the transduction of different CAR vectors. The performance of the process using healthy donor and a patient sample was evaluated. Functional assessment was performed of the CD19/22 and CD22 CAR T-cells using killing assays against the NALM6 tumor cell line and cytokine secretion analysis. Finally, gene expression of the transduced T-cells was examined to determine if there were any major changes that may have occurred as a result of the spinoculation process. Results The process of spinoculation lead to significant enhancement in gene transfer. Sepax spinoculation using a 1-h spin time showed comparable transduction efficiency to the bag spinoculation, and much greater than the static bag transduction method (83.4%, 72.8%, 35.7% n = 3). The performance of three different methods were consistent for all lentiviral vectors tested and no significant difference was observed when using starting cells from healthy donor versus a patient sample. Sepax spinoculation does not affect the function of the CAR T-cells against tumor cells, as these cells appeared to kill target cells equally well. Spinoculation also does not appear to affect gene expression patterns that are necessary for imparting function on the cell. Conclusions Closed system-bag spinoculation resulted in more efficient lymphocyte gene transfer than standard bag transductions without spinoculation. This method is effective for both retroviral and lentiviral vector gene transfer in lymphocytes and may be a feasible approach for gene transfer into other cell types including hematopoietic and myeloid progenitors. Sepax spinoculation further improved upon the process by offering an automated, closed system approach that significantly decreased hands-on time while also decreasing the risk of culture bag tears and microbial contamination.

2021 ◽  
Vol 8 ◽  
Hironobu Koseki ◽  
Shinya Sunagawa ◽  
Chieko Imai ◽  
Akihiko Yonekura ◽  
Umi Matsumura ◽  

Background: The operating theater is recognized to involve a high frequency of occupational blood and body fluid contacts.Objectives: This study aimed to visualize the production of blood and body fluid airborne particles by surgical procedures and to investigate risks of microbial contamination of the conjunctival membranes of surgical staff during orthopedic operations.Methods: Two physicians simulated total knee arthroplasty (TKA) and total hip arthroplasty (THA) in a bio-clean theater using model bones. The generation and behaviors of airborne particles were filmed using a fine particle visualization system, and numbers of airborne particles per 2.83 L of air were counted at the height of the operating and instrument tables. Each action was repeated five times, and particle counts were evaluated statistically.Results: Numerous airborne particles were dispersed to higher and wider areas while “cutting bones in TKA” and “striking and driving the cup component on the pelvic bone in THA” compared to other surgical procedures. The highest particle counts were detected while “cutting bones in TKA” under unidirectional laminar air flow.Discussion: These results provide a clearer image of the dispersion and distribution of airborne particles and identified higher-risk surgical procedures for microbial contamination of the conjunctival membranes. Surgical staff including surgeons, nurses, anesthesiologists, and visitors, should pay attention to and take measures against occupational infection particularly in high-risk surgical situations.

2022 ◽  
Vol 78 (01) ◽  
pp. 6616-2022

Feed microflora remains a very complex and still largely uncharacterized ecosystem. Given the wide range of potential sources of microbial contamination that may come into contact with feed, a variety of microorganisms, including pathogenic ones, can be expected. Microbiological contamination of feeds depends on environmental factors, which are a natural, primary source related to the microflora carried on feed materials and coming from soil, water and air. Microbial contamination may also emerge secondarily in the processing and distribution stages of feed, but also in the breeding stage, where feed may be contaminated by animals showing disease symptoms or asymptomatically. A wide variety of pathogenic microorganisms that are transmitted symptomatically or asymptomatically can cause economic losses to feed producers and farmers, and some of them due to their zoonotic nature can also pose a potential risk to consumers. New feed materials appear on the market, i.e. Insect Processed Animal Proteins, which are part of the strategy of replacing traditional protein sources. These materials are under investigation for their benefits as well as for microbiological safety. The aim of this review was to present the current knowledge on the main microbiological risk factors influencing the quality and safety of feed, as well as new analytical challenges related to the introduction of new feed materials.

Vanessa James ◽  
Hiral Panchal

Aim and Objective: The objective of the present study is to determine microbial contamination in fresh and packaged commercial fruit juices (including a combination of Aloe vera with fruit juices) available in the Ahmedabad city of Gujarat, India. Materials and Methods: Seventeen samples were collected from various parts of the city which includes 9 commercial fruit juice samples and 8 street vended fresh fruit juice samples. Samples were examined for Total plate count, Yeast and mould count, coliform count, Escherichia coli, Salmonella, Staphylococcus aureus, Shigella, Enterobacteriaceae, Listeria monocytogens and Vibrio Cholerae. Results: Commercial fruit juices do not exceed the FSSAI standards for fruit juices and are free of harmful pathogens making themsafe for human consumption. Street vended fresh fruit juice samples exceedthe FSSAI limit for Total Plate count, Yeast and mould count and Coliform count. Street vended Fresh fruit juices demonstratethepresence of Ecoli, Salmonella and Staphylococcus aureus in 75% (6/8) samples. Enterobacteriaceae were identified in street vended fruit juices which exceeds the FSSAI standard limit. Conclusion: The study demonstrates that commercial fruit juices were safe for human consumption but fresh juices showed significant microbial growth and harmful pathogens which must be controlled to ensure consumer’s safety and health. However regular monitoring of commercial and fresh fruit juices is recommended to avoid food borne illness resulting from pathogens encountered in the study.

2021 ◽  
Vol 23 (11) ◽  
pp. 297-312
Dalia Azher Ahmed ◽  
Zainab Zamel Khalaf ◽  
Hind Hussein Obaid ◽  

Thirty specimens of fresh white cheese, in different markets at different cities of Iraq were analyzed microbiologically. Isolates of E.coli that have been collected from the samples of cheese, were investigated. Capacity for biofilm producing was demonstrated by two method, Tissue culture plate method (TCP) and agar (CRA). After that, antibiofilm activity of lime extract and LiO2NPs was studied as each one of them alone and then the synergistic effect was done by TCP method. The results showed that all E.coli isolates produce biofilm but in different degrees. The results also displayed that Lime extract and LiO2NPs had antibiofilm effect against E.coli when used alone and when the combination done between each one of these materials. In conclusion, it was observed that the specimens of fresh white cheese included in this study contained microbial contamination at a health-threatening level but elimination of this contamination can be done by using lime extract and LiO2NPs.

Jared Johnson ◽  
Brandon Selover ◽  
Chris Curtin ◽  
Joy Waite-Cusic

The aim of this study was to investigate the temporal stability of microbial contamination during Cheddar cheese production by examining patterns of non-starter bacteria in 60-day aged Cheddar collected from the start and end of 30 consecutive production days. Further, we explored the source of these temporal microbial variations by comparing microbial communities in the aged cheese to those on food contact surfaces from a piece of cheesemaking equipment previously identified as a major source of non-starter bacteria in the same processing environment. 16S rRNA metabarcoding and culture-based sequencing methods identified two Streptococcus sequence variants significantly associated with the end of the production day in both the aged cheese and the cheese processing environment. Closer inspection of these sequence variants in the aged cheese over the 40-day sampling period revealed sinusoidal-like fluctuations in their relative ratios, which appeared to coincide with the Lactococcus starter rotation schedule. These results demonstrate that the microbial composition of finished cheese can vary according to the timing of processing within a production day. Further, our results demonstrate that time-of-day microbial differences in cheese can result from bacterial growth on food contact surfaces and that the composition of these microbial differences is subject to change day-to-day and may be linked to routine changes in the Lactococcus starter culture. Importance. Long production schedules used in modern cheese manufacturing can create circumstances which support the growth of microorganisms in the cheese processing environment. This work demonstrates that this growth can lead to significant changes in the microbial quality of aged cheese produced later in the production day. Further, we demonstrate that the dominant bacteria associated with these microbial changes throughout production are subject to change between days and might be influenced by specific cheese manufacturing practices. These findings improve understanding of microbial contamination patterns in modern food manufacturing facilities, therefore improving our ability to develop strategies to minimize quality losses due to microbial spoilage.

2021 ◽  
Bhargavi Rane ◽  
Alison Lacombe ◽  
Jiewen Guan ◽  
David F. Bridges ◽  
Shyam Sablani ◽  

Nabih Abdulrahman Baeshen ◽  
Nagwa Thabet Elsharawy ◽  
Naseebh Nabih Baeshen ◽  
Mohammed Nabih Baeshen

Background: The study describes the comparison of different microbial load results of natural falling and dipping of the house fly (Musca domestica) in water and milk to investigate the possibilities of preventing the effect of the transferred pathogens from the house fly to our sources by pointing out the existence of antimicrobial factors within the house fly. Methods: Samples of house fly were collected from Jeddah and Makkah (Makkah region) and were directly transferred to the laboratory. Each house fly was packed in sterile test tubes. Each tube was opened oppositely to a larger test tube containing 10 ml of sterile tap water, and sterile water at pH 4.0 in other similar series of treatments to represent the reactions of stomach fluids. Later, the house flies were left for 20 seconds after reaching the water surface, and then cultured on different microbial media to evaluate the microbial load of the natural falling of the house fly. To evaluate the complete dipping of house flies in the water, two methods were tested by one complete dip for the flies for 20 seconds, and three times complete dipping for 20 seconds in water before evaluating the microbial load. The same methods were achieved on milk in a series of experiments and the microbial load was evaluated after the incubation at room temperature for three hours. Results: It was found that dipping treatments of house flies gave lower microbial contamination in water at pH 4.0 than neutral pH. The lower microbial load was also observed when dipping the house flies three times in water as compared to once dipping and natural falling treatments. It was also found that the complete dipping of house flies’ treatments in milk will reduce the microbial contamination as compared to natural falling treatments. Conclusion: The observed results support the presence of antimicrobial factors on the house fly.

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