Search results for: foodborne pathogen

Authors: Pedro Batista, André Vinha, Filipe Castelo, Bárbara Costa, Ricardo Sousa, Raquel Ricardo, André Pinto

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Background: Septic arthritis is a diagnosis that must be considered in any patient presenting with acute joint swelling and fever. Among the several risk factors for septic arthritis, such as age, rheumatoid arthritis, recent surgery, or skin infection, diabetes mellitus can sometimes be the main risk factor. Staphylococcus aureus is the most common pathogen isolated in septic arthritis; however, it is uncommon in monomicrobial necrotizing fasciitis. Objectives: A case report of concomitant septic arthritis and necrotizing fasciitis in a patient with undiagnosed diabetes based on clinical history. Study Design & Methods: We report a case of a 58-year-old Portuguese previously healthy man who presented to the emergency department with fever and left knee swelling and pain for two days. The blood work revealed ketonemia of 6.7 mmol/L and glycemia of 496 mg/dL. The vital signs were significant for a temperature of 38.5 ºC and 123 bpm of heart rate. The left knee had edema and inflammatory signs. Computed tomography of the left knee showed diffuse edema of the subcutaneous cellular tissue and soft tissue air bubbles. A diagnosis of septic arthritis and necrotising fasciitis was made. He was taken to the operating room for surgical debridement. The samples collected intraoperatively were sent for microbiological analysis, revealing infection by multi-sensitive Staphylococcus aureus. Given this result, the empiric flucloxacillin (500 mg IV) and clindamycin (1000 mg IV) were maintained for 3 weeks. On the seventh day of hospitalization, there was a significant improvement in subcutaneous and musculoskeletal tissues. After two weeks of hospitalization, there was no purulent content and partial closure of the wounds was possible. After 3 weeks, he was switched to oral antibiotics (flucloxacillin 500 mg). A week later, a urinary infection by Pseudomonas aeruginosa was diagnosed and ciprofloxacin 500 mg was administered for 7 days without complications. After 30 days of hospital admission, the patient was discharged home and recovered. Results: The final diagnosis of concomitant septic arthritis and necrotizing fasciitis was made based on the imaging findings, surgical exploration and microbiological tests results. Conclusions: Early antibiotic administration and surgical debridement are key in the management of septic arthritis and necrotizing fasciitis. Furthermore, risk factors control (euglycemic blood glucose levels) must always be taken into account given the crucial role in the patient's recovery.

Keywords: septic arthritis, Necrotizing fasciitis, diabetes, Staphylococcus Aureus

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Authors: Jose R. Garcia, Lexi Frankel, Amalia Ardeljan, Sergio Medina, Ali Yasback, Omar Rashid

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Background: Helicobacter pylori (H. pylori) is a gram-negative, spiral-shaped bacterium that affects nearly half of the population worldwide and humans serve as the principal reservoir. Infection rates usually follow an inverse relationship with hygiene practices and are higher in developing countries than developed countries. Incidence varies significantly by geographic area, race, ethnicity, age, and socioeconomic status. H. pylori is primarily associated with conditions of the gastrointestinal tract such as atrophic gastritis and duodenal peptic ulcers. Infection is also associated with an increased risk of carcinogenesis as there is evidence to show that H. pylori infection may lead to gastric adenocarcinoma and mucosa-associated lymphoid tissue (MALT) lymphoma. It is suggested that H. pylori infection may be considered as a systemic condition, leading to various novel associations with several different neoplasms such as colorectal cancer, pancreatic cancer, and lung cancer, although further research is needed. Emerging evidence suggests that H. pylori infection may offer protective effects against Mycobacterium tuberculosis as a result of non-specific induction of interferon- γ (IFN- γ). Similar methods of enhanced immunity may affect the development of bronchogenic carcinoma due to the antiproliferative, pro-apoptotic and cytostatic functions of IFN- γ. The purpose of this study was to evaluate the correlation between Helicobacter pylori infection and the incidence of bronchogenic carcinoma. Methods: The data was provided by a Health Insurance Portability and Accountability Act (HIPAA) compliant national database to evaluate the patients infected versus patients not infected with H. pylori using ICD-10 and ICD-9 codes. Access to the database was granted by the Holy Cross Health, Fort Lauderdale for the purpose of academic research. Standard statistical methods were used. Results:-Between January 2010 and December 2019, the query was analyzed and resulted in 163,224 in both the infected and control group, respectively. The two groups were matched by age range and CCI score. The incidence of bronchogenic carcinoma was 1.853% with 3,024 patients in the H. pylori group compared to 4.785% with 7,810 patients in the control group. The difference was statistically significant (p < 2.22x10-16) with an odds ratio of 0.367 (0.353 - 0.383) with a confidence interval of 95%. The two groups were matched by treatment and incidence of cancer, which resulted in a total of 101,739 patients analyzed after this match. The incidence of bronchogenic carcinoma was 1.929% with 1,962 patients in the H. pylori and treatment group compared to 4.618% with 4,698 patients in the control group with treatment. The difference was statistically significant (p < 2.22x10-16) with an odds ratio of 0.403 (0.383 - 0.425) with a confidence interval of 95%.

Keywords: bronchogenic carcinoma, helicobacter pylori, lung cancer, pathogen-associated molecular patterns

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Authors: B. T. Naveen Kumar, Anuj Tyagi, Niraj Kumar Singh, Visanu Boonyawiwat, A. H. Shanthanagouda, Orawan Boodde, K. M. Shankar, Prakash Patil, Shubhkaramjeet Kaur

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Early mortality syndrome (EMS)/Acute Hepatopancreatic Necrosis Disease (AHPND) has emerged as a major obstacle for the shrimp farming around the world. It is caused by a strain of Vibrio parahaemolyticus. The possible preventive and control measure is, early and rapid detection of the pathogen in the broodstock, post-larvae and monitoring the shrimp during the culture period. Polymerase chain reaction (PCR) based early detection methods are good, but they are costly, time taking and requires a sophisticated laboratory. The present study was conducted to develop a simple, sensitive and rapid diagnostic farmer level kit for the reliable detection of AHPND in shrimp. A panel of monoclonal antibodies (MAbs) were raised against the recombinant Pir B protein (rPirB). First, an immunodot was developed by using MAbs G3B8 and Mab G3H2 which showed specific reactivity to purified r-PirB protein with no cross-reactivity to other shrimp bacterial pathogens (AHPND free Vibrio parahaemolyticus (Indian strains), V. anguillarum, WSSV, Aeromonas hydrophila, and Aphanomyces invadans). Immunodot developed using Mab G3B8 is more sensitive than that with the Mab G3H2. However, immunodot takes almost 2.5 hours to complete with several hands-on steps. Therefore, the flow-through assay (FTA) was developed by using a plastic cassette containing the nitrocellulose membrane with absorbing pads below. The sample was dotted in the test zone on the nitrocellulose membrane followed by continuos addition of five solutions in the order of i) blocking buffer (BSA) ii) primary antibody (MAb) iii) washing Solution iv) secondary antibody and v) chromogen substrate (TMB) clear purple dots against a white background were considered as positive reactions. The FTA developed using MAbG3B8 is more sensitive than that with MAb G3H2. In FTA the two MAbs showed specific reactivity to purified r-PirB protein and not to other shrimp bacterial pathogens. The FTA is simple to farmer/field level, sensitive and rapid requiring only 8-10 min for completion. Tests can be developed to kits, which will be ideal for use in biosecurity, for the first line of screening (at the port or pond site) and during monitoring and surveillance programmes overall for the good management practices to reduce the risk of the disease.

Keywords: acute hepatopancreatic necrosis disease, AHPND, flow-through assay, FTA, farmer level, immunodot, pond site, shrimp

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Authors: N. Demertzis, J. L. Bowen

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Sepsis is a complex and rapidly evolving condition, resulting from uncontrolled prolonged activation of host immune system due to pathogenic insult. The aim of this study is the development of a multiplex electrochemical sensing platform, capable of detecting both pathogen associated and host immune markers to enable the rapid and definitive diagnosis of sepsis. A combination of aptamers and molecular imprinting approaches have been employed to generate sensing systems for lipopolysaccharide (LPS), c-reactive protein (CRP) and procalcitonin (PCT). Gold working electrodes were mechanically polished and electrochemically cleaned with 0.1 M sulphuric acid using cyclic voltammetry (CV). Following activation, a self-assembled monolayer (SAM) was generated, by incubating the electrodes with a thiolated anti-LPS aptamer / dithiodibutiric acid (DTBA) mixture (1:20). 3-aminophenylboronic acid (3-APBA) in combination with the anti-LPS aptamer was used for the development of the hybrid molecularly imprinted sensor (apta-MIP). Aptasensors, targeting PCT and CRP were also fabricated, following the same approach as in the case of LPS, with mercaptohexanol (MCH) replacing DTBA. In the case of the CRP aptasensor, the SAM was formed following incubation of a 1:1 aptamer: MCH mixture. However, in the case of PCT, the SAM was formed with the aptamer itself, with subsequent backfilling with 1 μM MCH. The binding performance of all systems has been evaluated using electrochemical impedance spectroscopy. The apta-MIP’s polymer thickness is controlled by varying the number of electropolymerisation cycles. In the ideal number of polymerisation cycles, the polymer must cover the electrode surface and create a binding pocket around LPS and its aptamer binding site. Less polymerisation cycles will create a hybrid system which resembles an aptasensor, while more cycles will be able to cover the complex and demonstrate a bulk polymer-like behaviour. Both aptasensor and apta-MIP were challenged with LPS and compared to conventional imprinted (absence of aptamer from the binding site, polymer formed in presence of LPS) and non-imprinted polymers (NIPS, absence of LPS whilst hybrid polymer is formed). A stable LPS aptasensor, capable of detecting down to 5 pg/ml of LPS was generated. The apparent Kd of the system was estimated at 17 pM, with a Bmax of approximately 50 pM. The aptasensor demonstrated high specificity to LPS. The apta-MIP demonstrated superior recognition properties with a limit of detection of 1 fg/ml and a Bmax of 100 pg/ml. The CRP and PCT aptasensors were both able to detect down to 5 pg/ml. Whilst full binding performance is currently being evaluated, there is none of the sensors demonstrate cross-reactivity towards LPS, CRP or PCT. In conclusion, stable aptasensors capable of detecting LPS, PCT and CRP at low concentrations have been generated. The realisation of a multiplex panel such as described herein, will effectively contribute to the rapid, personalised diagnosis of sepsis.

Keywords: aptamer, electrochemical impedance spectroscopy, molecularly imprinted polymers, sepsis

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Authors: Nabeeha Hassan Abdel Jalil, Robert Barigye, Hamda Al Alawi, Afra Al Dhaheri, Fatma Graiban Al Muhairi, Maryam Al Khateri, Nouf Al Alalawi, Susan Olet, Khaja Mohteshamuddin, Ahmad Al Aiyan, Mohamed Elfatih Hamad

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A serological survey was carried out to determine the seroprevalence of Brucella spp, Coxiella burnetii, Chlamydophila abortus, and Toxoplasma gondii in sheep and goat herds in the UAE. A total of 915 blood samples [n= 222, [sheep]; n= 215, [goats]) were collected from livestock farms in the Emirates of Abu Dhabi, Dubai, Sharjah and Ras Al-Khaimah (RAK). An additional 478 samples (n= 244, [sheep]; n= 234, (goats]) were collected from the Al Ain livestock central market and tested by indirect ELISA for pathogen-specific antibodies with the Brucella antibodies being further corroborated by the Rose-Bengal agglutination test. Seropositivity for the four pathogens is variably documented in sheep and goats from the study area. Respectively, the overall livestock farm prevalence rates for Brucella spp, C. burnetii, C. abortus, and T. gondii were 2.7%, 27.9%, 8.1%, and 16.7% for sheep, and 0.0%, 31.6%, 9.3%, and 5.1% for goats. Additionally, the seroprevalence rates Brucella spp, C. burnetii, C. abortus, and T. gondii in samples from the livestock market were 7.4%, 21.7%, 16.4%, and 7.0% for sheep, and 0.9%, 32.5%, 19.2%, and 11.1% for goats respectively. Overall, sheep had 12.59 more chances than goats of testing seropositive for Brucella spp (OR, 12.59 [95% CI 2.96-53.6]) but less likely to be positive for C. burnetii-antibodies (OR, 0.73 [95% CI 0.54-0.97]). Notably, the differences in the seroprevalence rates of C. abortus and T. gondii in sheep and goats were not statistically significant (p > 0.0500). The present data indicate that all the four study pathogens are present in sheep and goat populations in the UAE where coxiellosis is apparently the most seroprevalent followed by chlamydophilosis, toxoplasmosis, and brucellosis. While sheep from the livestock market were more likely than those from farms to be Brucella-seropositive than those, the overall exposure risk of C. burnetii appears to be greater for goats than sheep. As more animals from the livestock market were more likely to be seropositive to Chlamydophila spp, it is possible that under the UAE animal production conditions, at least, coxiellosis and chlamydophilosis are more likely to increase the culling rate of domesticated small ruminants than toxoplasmosis and brucellosis. While anecdotal reports have previously insinuated that brucellosis may be a significant animal health risk in the UAE, the present data suggest C. burnetii, C. abortus and T. gondii to be more significant pathogens of sheep and goats in the country. Despite this possibility, the extent to which these pathogens may nationally be contributing to reproductive failure in sheep and goat herds is not known and needs to be investigated. Potentially, these agents may also carry a potentially zoonotic risk that needs to be investigated in risk groups like farm workers, and slaughter house personnel. An ongoing study is evaluating the seroprevalence of bovine coxiellosis in the Emirate of Abu Dhabi and the data thereof will further elucidate on the broader epidemiological dynamics of the disease in the national herd.

Keywords: Brucella spp, Chlamydophila abortus, goat, sheep, Toxoplasma gondii, UAE

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Authors: Suman Som, Abhijit Maity, Sunil B. Daschakraborty, Sujit Chaudhuri, Manik Pradhan

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Background: Helicobacter pylori is a common and important human pathogen and the primary cause of peptic ulcer disease and gastric cancer. Currently H. pylori infection is detected by both invasive and non-invasive way but the diagnostic accuracy is not up to the mark. Aim: To set up an optimal diagnostic cut-off value of 13C-Urea Breath Test to detect H. pylori infection and evaluate a novel c-PDR methodology to overcome of inconclusive grey zone. Materials and Methods: All 83 subjects first underwent upper-gastrointestinal endoscopy followed by rapid urease test and histopathology and depending on these results; we classified 49 subjects as H. pylori positive and 34 negative. After an overnight, fast patients are taken 4 gm of citric acid in 200 ml water solution and 10 minute after ingestion of the test meal, a baseline exhaled breath sample was collected. Thereafter an oral dose of 75 mg 13C-Urea dissolved in 50 ml water was given and breath samples were collected upto 90 minute for 15 minute intervals and analysed by laser based high precisional cavity enhanced spectroscopy. Results: We studied the excretion kinetics of 13C isotope enrichment (expressed as δDOB13C ‰) of exhaled breath samples and found maximum enrichment around 30 minute of H. pylori positive patients, it is due to the acid mediated stimulated urease enzyme activity and maximum acidification happened within 30 minute but no such significant isotopic enrichment observed for H. pylori negative individuals. Using Receiver Operating Characteristic (ROC) curve an optimal diagnostic cut-off value, δDOB13C ‰ = 3.14 was determined at 30 minute exhibiting 89.16% accuracy. Now to overcome grey zone problem we explore percentage dose of 13C recovered per hour, i.e. 13C-PDR (%/hr) and cumulative percentage dose of 13C recovered, i.e. c-PDR (%) in exhaled breath samples for the present 13C-UBT. We further explored the diagnostic accuracy of 13C-UBT by constructing ROC curve using c-PDR (%) values and an optimal cut-off value was estimated to be c-PDR = 1.47 (%) at 60 minute, exhibiting 100 % diagnostic sensitivity , 100 % specificity and 100 % accuracy of 13C-UBT for detection of H. pylori infection. We also elucidate the gastric emptying process of present 13C-UBT for H. pylori positive patients. The maximal emptying rate found at 36 minute and half empting time of present 13C-UBT was found at 45 minute. Conclusions: The present study exhibiting the importance of c-PDR methodology to overcome of grey zone problem in 13C-UBT for accurate determination of infection without any risk of diagnostic errors and making it sufficiently robust and novel method for an accurate and fast non-invasive diagnosis of H. pylori infection for large scale screening purposes.

Keywords: 13C-Urea breath test, c-PDR methodology, grey zone, Helicobacter pylori

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Authors: Gunjan Srivastava, Shamila Kalia

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Dalbergia sissoo Roxb., (Family- Leguminosae; Subfamily- Papilionoideae), is an economically and ecologically important tree species having medicinal value. Of the rich complex of insect fauna, ten have been recognized as potential pests of nurseries and plantations. Present study was conducted to explore an effective ecofriendly control of Dichomeris eridantis Meyrick, an important defoliator pest of D. sissoo. Health and environmental concerns demanded devising a bio-intensive pest management strategy and employing ecofriendly measures. In the present laboratory bioassay two entomopathogenic fungi Acremonium perscinum and Beauveria bassiana were tested and compared for evaluating the efficacy of their seven different concentrations (besides control) against the 3rd, 4th and 5th instar larvae of D. eridantis, on the basis of mean percent mortality data recorded and tabulated for seven days after treatment application. Analysis showed that both treatments vary significantly among themselves. Also, variations amongst instars and duration with respect to their mortality were highly significant (p < .001). All their interactions were found to vary significantly. B. bassiana at 0.25x107 spores / ml spore concentration caused maximum mean percent mortality (62.38%) followed by mean percent mortality at its 0.25x106 spores / ml concentration (56.67%). Mean percent mortality at maximum spore concentration (0.054x107 spores / ml) and next highest spore concentration (0.054 x106 spores / ml) due to A. perscinum treatment were far less effective (mean percent mortality of 45.40% and 31.29%, respectively). At 168 hours mean percent mortality of larval instars due to both fungal treatment applications reached its maximum (52.99%) whereas, at 24 hours mean percent mortality remained least (5.70%). In both cases, treatments were most effective against 3rd instar larvae and least effective against 5th instar larvae. A comparative acccount of efficacy of B. bassiana and A. perscinum on the 3rd, 4th and 5th instar larvae of D. eridantis on 5th, 6th and 7th post treatment observation days after their application, on the basis of their median lethal concentrations (LC50) proved B. bassiana to be more potential microbial pathogen of the two fungal microbes, for all the three instars (3rd, 4th and 5th) of D. eridantis, on all the three days (5th, 6th and 7th post observation days after application of both treatments). Percent mortality of D. eridantis increased in a dose dependent manner. Koch’s Postulates tested positive, thus confirming the pathogenicity of B. bassiana against the larval instars of D. eridantis. LC90 values of 0.280x1011 spores/ml, 0.301x108 spores/ml and 0.262x108 spores/ml concentrations of B. bassiana were standardized which can effectively cause mortality of all the larval instars of D. eridantis in the field after 5th, 6th and 7th day of their application, respectively. Therefore, these concentrations can be safely used in nurseries as well as plantations of D. sissoo for effective control of D. eridantis larvae.

Keywords: Acremonium perscinum, Beauveria bassiana, Dalbergia sissoo, Dichomeris eridantis

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Authors: Lena Lapidot, Amir Amnon, Rita Nosenko, Veitsman Ella, Cohen-Ezra Oranit, Davidov Yana, Segev Shlomo, Koren Omry, Safran Michal, Ben-Ari Ziv

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Introduction: Hepatocellular carcinoma (HCC) is one of the leading causes of cancer related mortality worldwide. Liver Cirrhosis is the main predisposing risk factor for the development of HCC. The factor(s) influencing disease progression from Cirrhosis to HCC remain unknown. Gut microbiota has recently emerged as a major player in different liver diseases, however its association with HCC is still a mystery. Moreover, there might be an important association between the gut microbiota, nutrition, life style and the progression of Cirrhosis and HCC. The aim of our study was to characterize the gut microbial signature in association with life style and nutrition of patients with Cirrhosis, HCC-Cirrhosis and healthy controls. Design: Stool samples were collected from 95 individuals (30 patients with HCC, 38 patients with Cirrhosis and 27 age, gender and BMI-matched healthy volunteers). All participants answered lifestyle and Food Frequency Questionnaires. 16S rRNA sequencing of fecal DNA was performed (MiSeq Illumina). Results: There was a significant decrease in alpha diversity in patients with Cirrhosis (qvalue=0.033) and in patients with HCC-Cirrhosis (qvalue=0.032) compared to healthy controls. The microbiota of patients with HCC-cirrhosis compared to patients with Cirrhosis, was characterized by a significant overrepresentation of Clostridium (pvalue=0.024) and CF231 (pvalue=0.010) and lower expression of Alphaproteobacteria (pvalue=0.039) and Verrucomicrobia (pvalue=0.036) in several taxonomic levels: Verrucomicrobiae, Verrucomicrobiales, Verrucomicrobiaceae and the genus Akkermansia (pvalue=0.039). Furthermore, we performed an analysis of predicted metabolic pathways (Kegg level 2) that resulted in a significant decrease in the diversity of metabolic pathways in patients with HCC-Cirrhosis (qvalue=0.015) compared to controls, one of which was amino acid metabolism. Furthermore, investigating the life style and nutrition habits of patients with HCC-Cirrhosis, we found significant correlations between intake of artificial sweeteners and Verrucomicrobia (qvalue=0.12), High sugar intake and Synergistetes (qvalue=0.021) and High BMI and the pathogen Campylobacter (qvalue=0.066). Furthermore, overweight in patients with HCC-Cirrhosis modified bacterial diversity (qvalue=0.023) and composition (qvalue=0.033). Conclusions: To the best of the our knowledge, we present the first report of the gut microbial composition in patients with HCC-Cirrhosis, compared with Cirrhotic patients and healthy controls. We have demonstrated in our study that there are significant differences in the gut microbiome of patients with HCC-cirrhosis compared to Cirrhotic patients and healthy controls. Our findings are even more pronounced because the significantly increased bacteria Clostridium and CF231 in HCC-Cirrhosis weren't influenced by diet and lifestyle, implying this change is due to the development of HCC. Further studies are needed to confirm these findings and assess causality.

Keywords: Cirrhosis, Hepatocellular carcinoma, life style, liver disease, microbiome, nutrition

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Authors: M. I. Ribah, M. Jibir, Y. A. Bashar, S. S. Manga

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Food safety is a significant and growing public health problem in the world and Nigeria as a developing country, since food-borne diseases are important contributors to the huge burden of sickness and death of humans. In Nigeria, traditional ready-to-eat meat products (RTE-MPs) like balangu, tsire, guru and dried meat products like kilishi, dambun nama, banda, were reported to be highly appreciated because of their eating qualities. The consumption of these products was considered as safe due to the treatments that are usually involved during their production process. However, during processing and handling, the products could be contaminated by pathogens that could cause food poisoning. Therefore, a hazard identification for pathogenic bacteria on some traditional RTE-MPs was conducted in Kebbi and Sokoto States, Nigeria. A total of 116 RTE-MPs (balangu-38, kilishi-39 and tsire-39) samples were obtained from retail outlets and analyzed using standard cultural microbiological procedures in general and selective enrichment media to isolate the target pathogens. A six-fold serial dilution was prepared and using the pour plating method, colonies were counted. Serial dilutions were selected based on the prepared pre-labeled Petri dishes for each sample. A volume of 10-12 ml of molten Nutrient agar cooled to 42-45°C was poured into each Petri dish and 1 ml each from dilutions of 102, 104 and 106 for every sample was respectively poured on a pre-labeled Petri plate after which colonies were counted. The isolated pathogens were identified and confirmed after series of biochemical tests. Frequencies and percentages were used to describe the presence of pathogens. The General Linear Model was used to analyze data on pathogen presence according to RTE-MPs and means were separated using the Tukey test at 0.05 confidence level. Of the 116 RTE-MPs samples collected, 35 (30.17%) samples were found to be contaminated with some tested pathogens. Prevalence results showed that Escherichia coli, salmonella and Staphylococcus aureus were present in the samples. Mean total bacterial count was 23.82×106 cfu/g. The frequency of individual pathogens isolated was; Staphylococcus aureus 18 (15.51%), Escherichia coli 12 (10.34%) and Salmonella 5 (4.31%). Also, among the RTE-MPs tested, the total bacterial counts were found to differ significantly (P < 0.05), with 1.81, 2.41 and 2.9×104 cfu/g for tsire, kilishi, and balangu, respectively. The study concluded that the presence of pathogenic bacteria in balangu could pose grave health risks to consumers, and hence, recommended good manufacturing practices in the production of balangu to improve the products’ safety.

Keywords: ready-to-eat meat products, retail outlets, public health, safety assessment

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Authors: Richa Sharma, Uma Nahar, Sadhna Sharma, Indu Verma

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Counteracting the deadly pathogen Mycobacterium tuberculosis (M. tb) effectively is still a global challenge. Scrutinizing alternative weapons like antimicrobial peptides to strengthen existing tuberculosis artillery is urgently required. Considering the antimycobacterial potential of Human Beta Defensin 1 (HBD-1) along with isoniazid, the present study was designed to explore the ability of HBD-1 to act against active and dormant M. tb. HBD-1 was screened in silico using antimicrobial peptide prediction servers to identify its short antimicrobial motif. The activity of both HBD-1 and its selected motif (Pep B) was determined at different concentrations against actively growing M. tb in vitro and ex vivo in monocyte derived macrophages (MDMs). Log phase M. tb was grown along with HBD-1 and Pep B for 7 days. M. tb infected MDMs were treated with HBD-1 and Pep B for 72 hours. Thereafter, colony forming unit (CFU) enumeration was performed to determine activity of both peptides against actively growing in vitro and intracellular M. tb. The dormant M. tb models were prepared by following two approaches and treated with different concentrations of HBD-1 and Pep B. Firstly, 20-22 days old M. tbH37Rv was grown in potassium deficient Sauton media for 35 days. The presence of dormant bacilli was confirmed by Nile red staining. Dormant bacilli were further treated with rifampicin, isoniazid, HBD-1 and its motif for 7 days. The effect of both peptides on latent bacilli was assessed by colony forming units (CFU) and most probable number (MPN) enumeration. Secondly, human PBMC granuloma model was prepared by infecting PBMCs seeded on collagen matrix with M. tb(MOI 0.1) for 10 days. Histopathology was done to confirm granuloma formation. The granuloma thus formed was incubated for 72 hours with rifampicin, HBD-1 and Pep B individually. Difference in bacillary load was determined by CFU enumeration. The minimum inhibitory concentrations of HBD-1 and Pep B restricting growth of mycobacteria in vitro were 2μg/ml and 20μg/ml respectively. The intracellular mycobacterial load was reduced significantly by HBD-1 and Pep B at 1μg/ml and 5μg/ml respectively. Nile red positive bacterial population, high MPN/ low CFU count and tolerance to isoniazid, confirmed the formation of potassium deficienybaseddormancy model. HBD-1 (8μg/ml) showed 96% and 99% killing and Pep B (40μg/ml) lowered dormant bacillary load by 68.89% and 92.49% based on CFU and MPN enumeration respectively. Further, H&E stained aggregates of macrophages and lymphocytes, acid fast bacilli surrounded by cellular aggregates and rifampicin resistance, indicated the formation of human granuloma dormancy model. HBD-1 (8μg/ml) led to 81.3% reduction in CFU whereas its motif Pep B (40μg/ml) showed only 54.66% decrease in bacterial load inside granuloma. Thus, the present study indicated that HBD-1 and its motif are effective antimicrobial players against both actively growing and dormant M. tb. They should be further explored to tap their potential to design a powerful weapon for combating tuberculosis.

Keywords: antimicrobial peptides, dormant, human beta defensin 1, tuberculosis

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Authors: Reena Aggarwal, Neha Kestwal

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The Himalayan Nettle (Girardinia diversifolia) has been used for centuries as fibre and food source by Himalayan communities. Himalayan Nettle is a natural cellulosic fibre that can be handled in the same way as other cellulosic fibres. The Uttarakhand Bamboo and Fibre Development Board based in Uttarakhand, India is working extensively with the nettle fibre to explore the potential of nettle for textile production in the region. The fiber is a potential resource for rural enterprise development for some high altitude pockets of the state and traditionally the plant fibre is used for making domestic products like ropes and sacks. Himalayan Nettle is an unconventional natural fiber with functional characteristics of shrink resistance, degree of pathogen and fire resistance and can blend nicely with other fibres. Most importantly, they generate mainly organic wastes and leave residues that are 100% biodegradable. The fabrics may potentially be reused or re-manufactured and can also be used as a source of cellulose feedstock for regenerated cellulosic products. Being naturally bio- degradable, the fibre can be composted if required. Though a lot of research activities and training are directed towards fibre extraction and processing techniques in different craft clusters villagers of different clusters of Uttarkashi, Chamoli and Bageshwar of Uttarakhand like retting and Degumming process, very little is been done to analyse the crucial properties of nettle fiber like shrinkage and wash fastness. These properties are very crucial to obtain desired quality of fibre for further processing of yarn making and weaving and in developing these fibers into fine saleable products. This research therefore is focused towards various on-field experiments which were focused on shrinkage properties conducted on cotton, nettle and cotton/nettle blended yarn samples. The objective of the study was to analyze the scope of the blended fiber for developing into wearable fabrics. For the study, after conducting the initial fiber length and fineness testing, cotton and nettle fibers were mixed in 60:40 ratio and five varieties of yarns were spun in open end spinning mill having yarn count of 3s, 5s, 6s, 7s and 8s. Samples of 100% Nettle 100% cotton fibers in 8s count were also developed for the study. All the six varieties of yarns were tested with shrinkage test and results were critically analyzed as per ASTM method D2259. It was observed that 100% Nettle has a least shrinkage of 3.36% while pure cotton has shrinkage approx. 13.6%. Yarns made of 100% Cotton exhibits four times more shrinkage than 100% Nettle. The results also show that cotton and Nettle blended yarn exhibit lower shrinkage than 100% cotton yarn. It was thus concluded that as the ratio of nettle increases in the samples, the shrinkage decreases in the samples. These results are very crucial for Uttarakhand people who want to commercially exploit the abundant nettle fiber for generating sustainable employment.

Keywords: Himalayan nettle, sustainable, shrinkage, blending

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Authors: Syed Asif Hassan, Tabrej Khan

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Tuberculosis (TB) caused by bacillus Mycobacterium tuberculosis (Mtb) continues to take a disturbing toll on human life and healthcare facility worldwide. The global burden of TB remains enormous. The alarming rise of multi-drug resistant strains of Mycobacterium tuberculosis calls for an increase in research efforts towards the development of new target specific therapeutics against diverse strains of M. tuberculosis. Therefore, the discovery of new molecular scaffolds targeting new drug sites should be a priority for a workable plan for fighting resistance in Mycobacterium tuberculosis (Mtb). Mtb non-acylated lipoprotein LprG (Rv1411c) has a Toll-like receptor 2 (TLR2) agonist actions that depend on its association with triacylated glycolipids binding specifically with the hydrophobic pocket of Mtb LprG lipoprotein. The detection of a glycolipid carrier function has important implications for the role of LprG in Mycobacterial physiology and virulence. Therefore, considering the pivotal role of glycolipids in mycobacterial physiology and host-pathogen interactions, designing competitive antagonist (chemotherapeutics) ligands that competitively bind to glycolipid binding domain in LprG lipoprotein, will lead to inhibition of tuberculosis infection in humans. In this study, a unified approach involving ligand-based virtual screening protocol USRCAT (Ultra Shape Recognition) software and molecular docking studies using Auto Dock Vina 1.1.2 using the X-ray crystal structure of Mtb LprG protein was implemented. The docking results were further confirmed by DSX (DrugScore eXtented), a robust program to evaluate the binding energy of ligands bound to the Ligand binding domain of the Mtb LprG lipoprotein. The ligand, which has the higher hypothetical affinity, also has greater negative value. Based on the USRCAT, Lipinski’s values and molecular docking results, [(2R)-2,3-di(hexadecanoyl oxy)propyl][(2S,3S,5S,6R)-3,4,5-trihydroxy-2,6-bis[[(2R,3S,4S,5R,6S)-3,4,5-trihydroxy-6 (hydroxymethyl)tetrahydropyran-2-yl]oxy]cyclohexyl] phosphate (XPX) was confirmed as a promising drug-like lead compound (antagonist) binding specifically to the hydrophobic domain of LprG protein with affinity greater than that of PIM2 (agonist of LprG protein) with a free binding energy of -9.98e+006 Kcal/mol and binding affinity of -132 Kcal/mol, respectively. A further, in vitro assay of this compound is required to establish its potency in inhibiting molecular evasion mechanism of MTB within the infected host macrophages. These results will certainly be helpful in future anti-TB drug discovery efforts against Multidrug-Resistance Tuberculosis (MDR-TB).

Keywords: antagonist, agonist, binding affinity, chemotherapeutics, drug-like, multi drug resistance tuberculosis (MDR-TB), RV1411c protein, toll-like receptor (TLR2)

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Authors: C. Watson, T. Glare, M. O'Callaghan, M. Hurst

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The endemic New Zealand grass grub (Costelytra giveni, Coleoptera: Scarabaeidae) is an economically significant grassland pest in New Zealand. Due to their impacts on production within the agricultural sector, one of New Zealand's primary industries, several methods are being used to either control or prevent the establishment of new grass grub populations in the pasture. One such method involves the use of a biopesticide based on the bacterium Serratia entomophila. This species is one of the causative agents of amber disease, a chronic disease of the larvae which results in death via septicaemia after approximately 2 to 3 months. The ability of S. entomophila to cause amber disease is dependant upon the presence of the amber disease associated plasmid (pADAP), which encodes for the key virulence determinants required for the establishment and maintenance of the disease. Following the collapse of grass grub populations within the soil, resulting from either natural population build-up or application of the bacteria, non-pathogenic plasmid-free Serratia strains begin to predominate within the soil. Whilst the interactions between S. entomophila and grass grub larvae are well studied, less information is known on the interactions between plasmid-bearing and plasmid-free strains, particularly the potential impact of these interactions upon the efficacy of an applied biopesticide. Using a range of constructed strains with antibiotic tags, in vitro (broth culture) and in vivo (soil and larvae) experiments were conducted using inoculants comprised of differing ratios of isogenic pathogenic and non-pathogenic Serratia strains, enabling the relative growth of pADAP+ and pADAP- strains under competition conditions to be assessed. In nutrient-rich, the non-pathogenic pADAP- strain outgrew the pathogenic pADAP+ strain by day 3 when inoculated in equal quantities, and by day 5 when applied as the minority inoculant, however, there was an overall gradual decline in the number of viable bacteria for both strains over a 7-day period. Similar results were obtained in additional experiments using the same strains and continuous broth cultures re-inoculated at 24-hour intervals, although in these cultures, the viable cell count did not diminish over the 7-day period. When the same ratios were assessed in soil microcosms with limited available nutrients, the strains remained relatively stable over a 2-month period. Additionally, in vivo grass grub co-infections assays using the same ratios of tagged Serratia strains revealed similar results to those observed in the soil, but there was also evidence of horizontal transfer of pADAP from the pathogenic to the non-pathogenic strain within the larval gut after a period of 4 days. Whilst the influence of competition is more apparent in broth cultures than within the soil or larvae, further testing is required to determine whether this competition between pathogenic and non-pathogenic Serratia strains has any influence on efficacy and disease progression, and how this may impact on the ability of S. entomophila to cause amber disease within grass grub larvae when applied as a biopesticide.

Keywords: biological control, entomopathogen, microbial ecology, New Zealand

Procedia PDF Downloads 156

Authors: Benjamin Castillo, Luis Pastenes, Fernando Cordova

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The pathogen growth in animal source foods is a common problem in the food industry, causing monetary losses due to the spoiling of products or food intoxication outbreaks in the community. In this sense, the quality of the product is reflected by the population of deteriorating agents present in it, which are mainly bacteria. The factors which are likely associated with freshness in animal source foods are temperature and processing, storage, and transport times. However, the level of deterioration of products depends, in turn, on the characteristics of the bacterial population, causing the decomposition or spoiling, such as pH level and toxins. Knowing the growth dynamics of the agents that are involved in product contamination allows the monitoring for more efficient processing. This means better quality and reasonable costs, along with a better estimation of necessary time and temperature intervals for transport and storage in order to preserve product quality. The objective of this project is to design a secondary model that allows measuring the impact on temperature bacterial growth and the competition for pH adequacy and release of bacteriocins in order to describe such phenomenon and, thus, estimate food product half-life with the least possible risk of deterioration or spoiling. In order to achieve this objective, the authors propose an analysis of a three-dimensional ordinary differential which includes; logistic bacterial growth extended by the inhibitory action of bacteriocins including the effect of the medium pH; change in the medium pH levels through an adaptation of the Luedeking-Piret kinetic model; Bacteriocin concentration modeled similarly to pH levels. These three dimensions are being influenced by the temperature at all times. Then, this differential system is expanded, taking into consideration the variable temperature and the concentration of pulsed bacteriocins, which represent characteristics inherent of the modeling, such as transport and storage, as well as the incorporation of substances that inhibit bacterial growth. The main results lead to the fact that temperature changes in an early stage of transport increased the bacterial population significantly more than if it had increased during the final stage. On the other hand, the incorporation of bacteriocins, as in other investigations, proved to be efficient in the short and medium-term since, although the population of bacteria decreased, once the bacteriocins were depleted or degraded over time, the bacteria eventually returned to their regular growth rate. The efficacy of the bacteriocins at low temperatures decreased slightly, which equates with the fact that their natural degradation rate also decreased. In summary, the implementation of the mathematical model allowed the simulation of a set of possible bacteria present in animal based products, along with their properties, in various transport and storage situations, which led us to state that for inhibiting bacterial growth, the optimum is complementary low constant temperatures and the initial use of bacteriocins.

Keywords: bacterial growth, bacteriocins, mathematical modelling, temperature

Procedia PDF Downloads 137

Authors: M. Shahin Alam, Satoru Takahashi, Mariko Itoh, Miyuki Komura, Mayuko Suzuki, Natthanan Sangsriratanakul, Kazuaki Takehara

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Cleaning and disinfection are key components of routine biosecurity in livestock farming and food processing industry. The usage of suitable disinfectants and their proper concentration are important factors for a successful biosecurity program. Disinfectants have optimum bactericidal and virucidal efficacies at temperatures above 20°C, but very few studies on application and effectiveness of disinfectants at low temperatures have been done. In the present study, the bactericidal efficacies of food additive grade calcium hydroxide (FdCa(OH)), quaternary ammonium compound (QAC) and their mixture, were investigated under different conditions, including time, organic materials (fetal bovine serum: FBS) and temperature, either in suspension or in carrier test. Salmonella Infantis and Escherichia coli, which are the most prevalent gram negative bacteria in commercial poultry housing and food processing industry, were used in this study. Initially, we evaluated these disinfectants at two different temperatures (4°C and room temperature (RT) (25°C ± 2°C)) and 7 contact times (0, 5 and 30 sec, 1, 3, 20 and 30 min), with suspension tests either in the presence or absence of 5% FBS. Secondly, we investigated the bactericidal efficacies of these disinfectants by carrier tests (rubber, stainless steel and plastic) at same temperatures and 4 contact times (30 sec, 1, 3, and 5 min). Then, we compared the bactericidal efficacies of each disinfectant within their mixtures, as follows. When QAC was diluted with redistilled water (dW2) at 1: 500 (QACx500) to obtain the final concentration of didecyl-dimethylammonium chloride (DDAC) of 200 ppm, it could inactivate Salmonella Infantis within 5 sec at RT either with or without 5% FBS in suspension test; however, at 4°C it required 30 min in presence of 5% FBS. FdCa(OH)2 solution alone could inactivate bacteria within 1 min both at RT and 4°C even with 5% FBS. While FdCa(OH)2 powder was added at final concentration 0.2% to QACx500 (Mix500), the mixture could inactivate bacteria within 30 sec and 5 sec, respectively, with or without 5% FBS at 4°C. The findings from the suspension test indicated that low temperature inhibited the bactericidal efficacy of QAC, whereas Mix500 was effective, regardless of short contact time and low temperature, even with 5% FBS. In the carrier test, single disinfectant required bit more time to inactivate bacteria on rubber and plastic surfaces than on stainless steel. However, Mix500 could inactivate S. Infantis on rubber, stainless steel and plastic surfaces within 30 sec and 1 min, respectively, at RT and 4°C; but, for E. coli, it required only 30 sec at both temperatures. So, synergistic effects were observed on different carriers at both temperatures. For a successful enhancement of biosecurity during winter, the disinfectants should be selected that could have short contact times with optimum efficacy against the target pathogen. The present study findings help farmers to make proper strategies for application of disinfectants in their livestock farming and food processing industry.

Keywords: carrier, food additive grade calcium hydroxide (FdCa(OH)₂), quaternary ammonium compound, synergistic effects

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Authors: Norihiro Kato, Yuriko Takayama

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Some gram-negative bacteria enable the simultaneous activation of gene expression involved in N-acylhomoserine lactone (AHL) dependent cell-to-cell communication system. Such regulatory system for the bacterial group behavior is termed as quorum sensing (QS) because a diffusible AHL signal can accumulate around the cell during the increase of the cell density and trigger activation of the sequential QS process. By blocking the QS, the expression of diverse genes related to infection, antibiotic production, and biofilm formation is inhibited. Conditioning of QS by regulation of the DNA-receptor-AHL interaction is a potential target for enhancing host defenses against pathogenicity. We focused on engineered application of transcriptional regulator SpnR produced in opportunistic human pathogen Serratia marcescens. The SpnR can interact with AHL signals at an N-terminal domain and also with a promoter region of a QS target gene at a C-terminal domain. As the initial process of the QS activation, the SpnR forms a complex with the AHL to enhance the expression of pig cluster; the SpnR normally acts as an activator for the expression of the QS-dependent gene. In this research, we attempt to artificially control QS by changing the role of SpnR. The QS-dependent prodigiosin production is expected to inhibit by externally added SpnR in the culture broth of AS-1 strain because the AHL concentration was kept below the threshold by AHL-SpnR complex formation. Maltose-binding protein (MBP)-tagged SpnR (MBP-SpnR) was overexpressed in Escherichia coli and purified using an affinity chromatography equipped with an amylose resin column. The specific interaction between AHL and MBP-SpnR was demonstrated by quartz crystal microbalance (QCM) sensor. AHL with amino end-group was coupled with COOH-terminated self-assembled monolayer prepared on a gold electrode of 27-MHz quartz crystal sensor using water-soluble carbodiimide. After the injection of MBP-SpnR into a cup-type sensor cell filled with the buffer solution, time course of resonant frequency change (ΔFs) was determined. A decrease of ΔFs clearly showed the uptake of MBP-SpnR onto the AHL-immobilized electrode. Furthermore, no binding affinity was observed after the heat-inactivation of MBP-SpnR at 80ºC. These results suggest that MBP-SpnR possesses a specific affinity for AHL. MBP-SpnR was added to the culture medium as an AHL trap to study inhibitory effects on intracellularly accumulated prodigiosin. With approximately 2 µM MBP-SpnR, the amount of prodigiosin induced was half that of the control without any additives. In conclusion, the function of SpnR could be switched by adding it to the cell culture. Exogenously added MBP-SpnR possesses high affinity for AHL derived from cells and acts as an inhibitor of AHL-mediated QS.

Keywords: intracellular signaling, microbial biotechnology, quorum sensing, transcriptional regulator

Procedia PDF Downloads 267

Authors: Mudasir Ahmad Syed, Raashid Ahmd Wani, Mashooq Ahmad Dar, Uneeb Urwat, Riaz Ahmad Shah, Nazir Ahmad Ganai

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Salmonella enterica serovar Typhimurium (Salmonella Typhimurium) is a primary avian pathogen responsible for severe intestinal pathology in younger chickens and economic losses. However, the Salmonella Typhimurium is also able to cause infection in humans, described by typhoid fever and acute gastro-intestinal disease. A study was conducted at days to investigate pathological, histopathological, haemato-biochemical, immunological and expression kinetics of NRAMP (natural resistance associated macrophage protein) gene family (NRAMP1 and NRAMP2) in broiler chickens following experimental infection of Salmonella Typhimurium at 0,1,3,5,7,9,11,13 and 15 days respectively. Infection was developed in birds through oral route at 2×108 CFU/ml. Clinical symptoms appeared 4 days post infection (dpi) and after one-week birds showed progressive weakness, anorexia, diarrhea and lowering of head. On postmortem examination, liver showed congestion, hemorrhage and necrotic foci on surface, while as spleen, lungs and intestines revealed congestion and hemorrhages. Histopathological alterations were principally observed in liver in second week post infection. Changes in liver comprised of congestion, areas of necrosis, reticular endothelial hyperplasia in association with mononuclear cell and heterophilic infiltration. Hematological studies confirm a significant decrease (P<0.05) in RBC count, Hb concentration and PCV. White blood cell count showed significant increase throughout the experimental study. An increase in heterophils was found up to 7dpi and a decreased pattern was observed afterwards. Initial lymphopenia followed by lymphocytosis was found in infected chicks. Biochemical studies showed a significant increase in glucose, AST and ALT concentration and a significant decrease (P<0.05) in total protein and albumin level in the infected group. Immunological studies showed higher titers of IgG in infected group as compared to control group. The real time gene expression of NRAMPI and NRAMP2 genes increased significantly (P<0.05) in infected group as compared to controls. The peak expression of NRAMP1 gene was seen in liver, spleen and caecum of infected birds at 3dpi, 5dpi and 7dpi respectively, while as peak expression of NRAMP2 gene in liver, spleen and caecum of infected chicken was seen at 9dpi, 5dpi and 9dpi respectively. This study has role in diagnostics and prognostics in the poultry industry for the detection of salmonella infections at early stages of poultry development.

Keywords: biochemistry, histopathology, NRAMP, poultry, real time expression, Salmonella Typhimurium

Procedia PDF Downloads 332

Authors: Abheyjit Singh, Kulwant Singh

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There is a gradual depletion of the water table in the earth's crust, and it is required to converse and reduce the scarcity of water. This is only done by rainwater harvesting, recycling of water and by judicially consumption/utilization of water and adopting unique treatment measures. Domestic waste is generated in residential areas, commercial settings, and institutions. Waste, in general, is unwanted, undesirable, and nevertheless an inevitable and inherent product of social, economic, and cultural life. In a cluster, a need-based system is formed where the project is designed for systematic analysis, collection of sewage from the cluster, treating it and then recycling it for multifarious work. The liquid waste may consist of Sanitary sewage/ Domestic waste, Industrial waste, Storm waste, or Mixed Waste. The sewage contains both suspended and dissolved particles, and the total amount of organic material is related to the strength of the sewage. The untreated domestic sanitary sewage has a BOD (Biochemical Oxygen Demand) of 200 mg/l. TSS (Total Suspended Solids) about 240 mg/l. Industrial Waste may have BOD and TSS values much higher than those of sanitary sewage. Another type of impurities of wastewater is plant nutrients, especially when there are compounds of nitrogen N phosphorus P in the sewage; raw sanitary contains approx. 35 mg/l Nitrogen and 10 mg/l of Phosphorus. Finally, the pathogen in the waste is expected to be proportional to the concentration of facial coliform bacteria. The coliform concentration in raw sanitary sewage is roughly 1 billion per liter. The system of sewage disposal technique has been universally applied to all conditions, which are the nature of soil formation, Availability of land, Quantity of Sewage to be disposed of, The degree of treatment and the relative cost of disposal technique. The adopted Thappar Model (India) has the following designed parameters consisting of a Screen Chamber, a Digestion Tank, a Skimming Tank, a Stabilization Tank, an Oxidation Pond and a Water Storage Pond. The screening Chamber is used to remove plastic and other solids, The Digestion Tank is designed as an anaerobic tank having a retention period of 8 hours, The Skimming Tank has an outlet that is kept 1 meter below the surface anaerobic condition at the bottom and also help in organic solid remover, Stabilization Tank is designed as primary settling tank, Oxidation Pond is a facultative pond having a depth of 1.5 meter, Storage Pond is designed as per the requirement. The cost of the Thappar model is Rs. 185 Lakh per 3,000 to 4,000 population, and the Area required is 1.5 Acre. The complete structure will linning as per the requirement. The annual maintenance will be Rs. 5 lakh per year. The project is useful for water conservation, silage water for irrigation, decrease of BOD and there will be no longer damage to community assets and economic loss to the farmer community by inundation. There will be a healthy and clean environment in the community.

Keywords: collection, treatment, utilization, economic

Procedia PDF Downloads 79

Authors: Sundru Manjulata Devi, Prakash M. Halami

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The immemorial use of fermented foods from vegetables, dairy and other biological sources are of great demand in India because of their health benefits. However, the diversity of Lactobacillus plantarum group (LPG) of vegetable origin has not been revealed yet, particularly with reference to their probiotic functionalities. In the present study, the different species of probiotic Lactobacillus plantarum group (LPG) i.e., L. plantarum subsp. plantarum MTCC 5422 (from fermented cereals), L. plantarum subsp. argentoratensis FG16 (from fermented bamboo shoot) and L. paraplantarum MTCC 9483 (from fermented gundruk) (as characterized by multiplex recA PCR assay) were considered to investigate their relative expression of MUB domains of mub gene (mucin binding protein) by Real time PCR. Initially, the allelic variation in the mub gene was assessed and found to encode three different variants (Type I, II and III). All the three types had 8, 9 and 10 MUB domains respectively (as analysed by Pfam database) and were found to be responsible for adhesion of bacteria to the host intestinal epithelial cells. These domains either get inserted or deleted during speciation or evolutionary events and lead to divergence. The reverse transcriptase qPCR analysis with mubLPF1+R1 primer pair supported variation in amplicon sizes with 300, 500 and 700 bp among different LPG strains. The relative expression of these MUB domains significantly unregulated in the presence of 1% mucin in overnight grown cultures. Simultaneously, the mub gene expressed efficiently by 7 fold in the culture L. paraplantarum MTCC 9483 with 10 MUB domains. An increase in the expression levels for L. plantarum subsp. plantarum MTCC 5422 and L. plantarum subsp. argentoratensis FG16 (MCC 2974) with 9 and 8 repetitive domains was around 4 and 2 fold, respectively. The detection and expression of an integrase (int) gene in the upstream region of mub gene reveals the excision and integration of these repetitive domains. Concurrently, an in vitro adhesion assay to mucin and exclusion of pathogens (such as Listeria monocytogenes and Micrococcus leuteus) was investigated and observed that the L. paraplantarum MTCC 9483 with more adhesion domains has more ability to adhere to mucin and inhibited the growth of pathogens. The production and expression of plantaricin-like bacteriocin (plnNC8 type) in MTCC 9483 suggests the pathogen inhibition. Hence, the expression of MUB domains can act as potential biomarkers in the screening of a novel probiotic LPG strain with adherence property. The present study provides a platform for an easy, rapid, less time consuming, low-cost methodology for the detection of potential probiotic bacteria. It was known that the traditional practices followed in the preparation of fermented bamboo shoots/gundruk/cereals of Indian foods contain different kinds of neutraceuticals for functional food and novel compounds with health promoting factors. In future, a detailed study of these food products can add more nutritive value, consumption and suitable for commercialization.

Keywords: adhesion gene, fermented foods, MUB domains, probiotics

Procedia PDF Downloads 271

Authors: Uraib Sharaha, Ahmad Salman, Eladio Rodriguez-Diaz, Elad Shufan, Klaris Riesenberg, Irving J. Bigio, Mahmoud Huleihel

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Antimicrobial drugs have played an indispensable role in controlling illness and death associated with infectious diseases in animals and humans. However, the increasing resistance of bacteria to a broad spectrum of commonly used antibiotics has become a global healthcare problem. Many antibiotics had lost their effectiveness since the beginning of the antibiotic era because many bacteria have adapted defenses against these antibiotics. Rapid determination of antimicrobial susceptibility of a clinical isolate is often crucial for the optimal antimicrobial therapy of infected patients and in many cases can save lives. The conventional methods for susceptibility testing require the isolation of the pathogen from a clinical specimen by culturing on the appropriate media (this culturing stage lasts 24 h-first culturing). Then, chosen colonies are grown on media containing antibiotic(s), using micro-diffusion discs (second culturing time is also 24 h) in order to determine its bacterial susceptibility. Other methods, genotyping methods, E-test and automated methods were also developed for testing antimicrobial susceptibility. Most of these methods are expensive and time-consuming. Fourier transform infrared (FTIR) microscopy is rapid, safe, effective and low cost method that was widely and successfully used in different studies for the identification of various biological samples including bacteria; nonetheless, its true potential in routine clinical diagnosis has not yet been established. The new modern infrared (IR) spectrometers with high spectral resolution enable measuring unprecedented biochemical information from cells at the molecular level. Moreover, the development of new bioinformatics analyses combined with IR spectroscopy becomes a powerful technique, which enables the detection of structural changes associated with resistivity. The main goal of this study is to evaluate the potential of the FTIR microscopy in tandem with machine learning algorithms for rapid and reliable identification of bacterial susceptibility to antibiotics in time span of few minutes. The UTI E.coli bacterial samples, which were identified at the species level by MALDI-TOF and examined for their susceptibility by the routine assay (micro-diffusion discs), are obtained from the bacteriology laboratories in Soroka University Medical Center (SUMC). These samples were examined by FTIR microscopy and analyzed by advanced statistical methods. Our results, based on 700 E.coli samples, were promising and showed that by using infrared spectroscopic technique together with multivariate analysis, it is possible to classify the tested bacteria into sensitive and resistant with success rate higher than 90% for eight different antibiotics. Based on these preliminary results, it is worthwhile to continue developing the FTIR microscopy technique as a rapid and reliable method for identification antibiotic susceptibility.

Keywords: antibiotics, E.coli, FTIR, multivariate analysis, susceptibility, UTI

Procedia PDF Downloads 174

Authors: Marina Sofia, Alexios Giannakopoulos, Antonia Touloudi, Dimitris C Chatzopoulos, Zoi Athanasakopoulou, Vassiliki Spyrou, Charalambos Billinis

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Fertility of dairy cattle can be affected by heat stress when the ambient temperature increases above 30°C and the relative humidity ranges from 35% to 50%. The present study was conducted on dairy cattle farms during summer months in Greece and aimed to identify the serological profile against pathogens that could affect fertility and to associate the positive serological results at herd level with temperature factors. A total of 323 serum samples were collected from clinically healthy dairy cows of 8 herds, located in Central and Northern Greece. ELISA tests were performed to detect antibodies against selected pathogens that affect fertility, namely Chlamydophila abortus, Coxiella burnetii, Neospora caninum, Toxoplasma gondii and Infectious Bovine Rhinotracheitis Virus (IBRV). Eleven climatic variables were derived from the WorldClim version 1.4. and ArcGIS V.10.1 software was used for analysis of the spatial information. Five different MaxEnt models were applied to associate the temperature variables with the locations of seropositive Chl. abortus, C. burnetii, N. caninum, T. gondii and IBRV herds (one for each pathogen). The logistic outputs were used for the interpretation of the results. ROC analyses were performed to evaluate the goodness of fit of the models’ predictions. Jackknife tests were used to identify the variables with a substantial contribution to each model. The seropositivity rates of pathogens varied among the 8 herds (0.85-4.76% for Chl. abortus, 4.76-62.71% for N. caninum, 3.8-43.47% for C. burnetii, 4.76-39.28% for T. gondii and 47.83-78.57% for IBRV). The variables of annual temperature range, mean diurnal range and maximum temperature of the warmest month gave a contribution to all five models. The regularized training gains, the training AUCs and the unregularized training gains were estimated. The mean diurnal range gave the highest gain when used in isolation and decreased the gain the most when it was omitted in the two models for seropositive Chl.abortus and IBRV herds. The annual temperature range increased the gain when used alone and decreased the gain the most when it was omitted in the models for seropositive C. burnetii, N. caninum and T. gondii herds. In conclusion, antibodies against Chl. abortus, C. burnetii, N. caninum, T. gondii and IBRV were detected in most herds suggesting circulation of pathogens that could cause infertility. The results of the spatial analyses demonstrated that the annual temperature range, mean diurnal range and maximum temperature of the warmest month could affect positively the possible pathogens’ presence. Acknowledgment: This research has been co‐financed by the European Regional Development Fund of the European Union and Greek national funds through the Operational Program Competitiveness, Entrepreneurship and Innovation, under the call RESEARCH–CREATE–INNOVATE (project code: T1EDK-01078).

Keywords: dairy cows, seropositivity, spatial analysis, temperature factors

Procedia PDF Downloads 200

Authors: Celine Shi Ying Lee, Jong Jian Lee

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Background: Infectious keratitis is one of the leading causes of blindness worldwide. Prompt treatment with effective medication will control the infection early, preventing corneal scarring and visual loss. fluoroquinolones ophthalmic medication is used because of its broad-spectrum properties, potency, good intraocular penetration, and low toxicity. The study aims to evaluate the treatment outcome of corneal ulcers using Levofloxacin 1.5% ophthalmic solution (LVFX) with adjuvant oral ciprofloxacin when indicated and apply this treatment strategy in primary health care as first-line treatment. Methods: Patients with infective corneal ulcer treated in an eye center were recruited. Inclusion criteria includes Corneal infection consistent with bacterial keratitis, single or multiple small corneal ulcers. Treatment regime: LVFX hourly for the first 2 days, 2 hourly from the 3rd day, and 3 hourly on the 5th day of review. Adjuvant oral ciprofloxacin 500mg BD was administered for 5 days if there were multiple corneal ulcers or when the location of the cornea ulcer was central or paracentral. Results: 47 subjects were recruited. There were 16 (34%) males and 31 (66%) females. 40 subjects (85%) were contact lens (CL) related to corneal ulcer, and 7 subjects (15%) were non-contact lens related. 42 subjects (89%) presented with one ulcer, of which 20 of them (48%) needed adjuvant therapy. 5 subjects presented with 2 or 3 ulcers, of which 3 needed adjuvant therapy. A total of 23 subjects (49%) was given adjuvant therapy (oral ciprofloxacin 500mg BD for 5 days).21 of them (91%) were CL related. All subjects recovered fully, and the average duration of treatment was 3.7 days, with 49% of the subjects resolved on the 3rd day, 38% on the 5thday of and 13% on the 7thday. All subjects showed symptoms of relief of pain, light-sensitivity, and redness on the 3rd day with full visual recovery post-treatment. No adverse drug reactions were recorded. Conclusion: Our treatment regime demonstrated good clinical outcome as first-line treatment for corneal ulcers. A corneal ulcer is a common eye condition in Singapore, mainly due to CL wear. Pseudomonas aeruginosa is the most frequent and potentially sight-threatening pathogen involved in CL related corneal ulcer. Coagulase-negative Staphylococci, Staphylococcus aureus, and Streptococcus Pneumoniae were seen in non-CL users. All these bacteria exhibit good sensitivity rates to ciprofloxacin and levofloxacin. It is therefore logical in our study to use LVFX Eyedrops and adjuvant ciprofloxacin oral antibiotics when indicated as first line treatment for most corneal ulcers. Our study of patients, both CL related and non-CL related, have shown good clinical response and full recovery using the above treatment strategy. There was also a full restoration of visual acuity in all the patients. Eye-trained primary Healthcare practitioners can consider adopting this treatment strategy as first line treatment in patients with corneal ulcers. This is relevant during the COVID pandemic, where hospitals are overwhelmed with patients and in regions with limited access to specialist eye care. This strategy would enable early treatment with better clinical outcome.

Keywords: corneal ulcer, levofloxacin hydrate, treatment strategy, ciprofloxacin

Procedia PDF Downloads 175

Authors: Abirami Baleswaran, Christel Couderc, Loubnah Belahcen, Jean Dayde, Hélène Tormo, Gwénaëlle Jard

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Cheeses are often reported to be spoiled by Pseudomonas spp., responsible for defects in appearance, texture, taste, and smell, leading to their non-marketing and even their destruction. Despite preventive actions, problems linked to Pseudomonas spp. are difficult to control by the lack of knowledge and control of these contaminants during the cheese manufacturing. Lactic goat cheese producers are not spared by this problem and are looking for solutions to decrease the number of spoiled cheeses. To explore different hypotheses, experiments are needed. However, cheese-making experiments at the pilot scale are expensive and time consuming. Thus, there is a real need to develop a miniature cheeses model system under controlled conditions. In a previous study, several miniature cheese models corresponding to different type of commercial cheeses have been developed for different purposes. The models were, for example, used to study the influence of milk, starters cultures, pathogen inhibiting additives, enzymatic reactions, microflora, freezing process on cheese. Nevertheless, no miniature model was described on the lactic goat cheese. The aim of this work was to develop a miniature cheese model system under controlled laboratory conditions which resembles commercial lactic goat cheese to study Pseudomonas spp. spoilage during the manufacturing and ripening process. First, a protocol for the preparation of miniature cheeses (3.5 times smaller than a commercial one) was designed based on the cheese factorymanufacturing process. The process was adapted from “Rocamadour” technology and involves maturation of pasteurized milk, coagulation, removal of whey by centrifugation, moulding, and ripening in a little scale cellar. Microbiological (total bacterial count, yeast, molds) and physicochemical (pH, saltinmoisture, moisture in fat-free)analyses were performed on four key stages of the process (before salting, after salting, 1st day of ripening, and end of ripening). Factory and miniature cheeses volatilomewere also obtained after full scan Sift-MS cheese analysis. Then, Pseudomonas spp. strains isolated from contaminated cheeses were selected on their origin, their ability to produce pigments, and their enzymatic activities (proteolytic, lecithinasic, and lipolytic). Factory and miniature curds were inoculated by spotting selected strains on the cheese surface. The expression of cheese spoilage was evaluated by counting the level of Pseudomonas spp. during the ripening and by visual observation and under UVlamp. The physicochemical and microbiological compositions of miniature cheeses permitted to assess that miniature process resembles factory process. As expected, differences involatilomes were observed, probably due to the fact that miniature cheeses are made usingpasteurized milk to better control the microbiological conditions and also because the little format of cheese induced probably a difference during the ripening even if the humidity and temperature in the cellar were quite similar. The spoilage expression of Pseudomonas spp. was observed in miniature and factory cheeses. It confirms that the proposed model is suitable for the preparation of miniature cheese specimens in the spoilage study of Pseudomonas spp. in lactic cheeses. This kind of model could be deployed for other applications and other type of cheese.

Keywords: cheese, miniature, model, pseudomonas spp, spoilage

Procedia PDF Downloads 134

Authors: Jone Ibarruri, Mikel Manso, Marta Cebrián

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A high amount of food is lost or discarded in the World every year. In addition, in the last decades, an increasing demand of new alternative and sustainable sources of proteins and other valuable compounds is being observed in the food and feeding sectors and, therefore, the use of food by-products as nutrients for these purposes sounds very interesting from the environmental and economical point of view. However, the direct use of discarded fruit and vegetables that present, in general, a low protein content is not interesting as feeding ingredient except if they are used as a source of fiber for ruminants. Especially in the case of aquaculture, several alternatives to the use of fish meal and other vegetable protein sources have been extensively explored due to the scarcity of fish stocks and the unsustainability of fishing for these purposes. Fish mortality is also of great concern in this sector as this problem highly reduces their economic feasibility. So, the development of new functional and natural ingredients that could reduce the need for vaccination is also of great interest. In this work, several fermentation tests were developed at lab scale using a selected mixture of fruit and vegetable discards from a wholesale market located in the Basque Country to increase their protein content and also to produce some bioactive extracts that could be used as additives in aquaculture. Fruit and vegetable mixtures (60/40 ww) were centrifugated for humidity reduction and crushed to 2-5 mm particle size. Samples were inoculated with a selected Rhizopus oryzae strain and fermented for 7 days in controlled conditions (humidity between 65 and 75% and 28ºC) in Petri plates (120 mm) by triplicate. Obtained results indicated that the final fermented product presented a twofold protein content (from 13 to 28% d.w). Fermented product was further processed to determine their possible functionality as a feed additive. Extraction tests were carried out to obtain an ethanolic extract (60:40 ethanol: water, v.v) and remaining biomass that also could present applications in food or feed sectors. The extract presented a polyphenol content of about 27 mg GAE/gr d.w with antioxidant activity of 8.4 mg TEAC/g d.w. Remining biomass is mainly composed of fiber (51%), protein (24%) and fat (10%). Extracts also presented antibacterial activity according to the results obtained in Agar Diffusion and to the Minimum Inhibitory Concentration (MIC) tests determined against several food and fish pathogen strains. In vitro, digestibility was also assessed to obtain preliminary information about the expected effect of extraction procedure on fermented product digestibility. First results indicated that remaining biomass after extraction doesn´t seem to improve digestibility in comparison to the initial fermented product. These preliminary results show that fermented fruit and vegetables can be a useful source of functional ingredients for aquaculture applications and a substitute of other protein sources in the feeding sector. Further validation will be also carried out through “in vivo” tests with trout and bass.

Keywords: fungal solid state fermentation, protein increase, functional extracts, feed ingredients

Procedia PDF Downloads 64

Authors: Jovana Grahovac, Ivana Pajcin, Natasa Lukic, Jelena Dodic, Aleksandar Jokic

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To obtain purified biomass to be used in the plant pathogen biocontrol or as soil biofertilizer, it is necessary to eliminate residual broth components at the end of the fermentation process. The main drawback of membrane separation techniques is permeate flux decline due to the membrane fouling. Fouling mitigation measures increase the pressure drop along membrane channel due to the increased resistance to flow of the feed suspension, thus increasing the hydraulic power drop. At the same time, these measures lead to an increase in the permeate flux due to the reduced resistance of the filtration cake on the membrane surface. Because of these opposing effects, the energy efficiency of fouling mitigation measures is limited, and the justification of its application is provided by information on a reducing specific energy consumption compared to a case without any measures employed. In this study, the influence of static mixer (Kenics) and air-sparging (two-phase flow) on reduction of specific energy consumption (ER) was investigated. Cultivation Bacillus velezensis was carried out in the 3-L bioreactor (Biostat® Aplus) containing 2 L working volume with two parallel Rushton turbines and without internal baffles. Cultivation was carried out at 28 °C on at 150 rpm with an aeration rate of 0.75 vvm during 96 h. The experiments were carried out in a conventional cross-flow microfiltration unit. During experiments, permeate and retentate were recycled back to the broth vessel to simulate continuous process. The single channel ceramic membrane (TAMI Deutschland) used had a nominal pore size 200 nm with the length of 250 mm and an inner/external diameter of 6/10 mm. The useful membrane channel surface was 4.33×10⁻³ m². Air sparging was brought by the pressurized air connected by a three-way valve to the feed tube by a simple T-connector without diffusor. The different approaches to flux improvement are compared in terms of energy consumption. Reduction of specific energy consumption compared to microfiltration without fouling mitigation is around 49% and 63%, for use of two-phase flow and a static mixer, respectively. In the case of a combination of these two fouling mitigation methods, ER is 60%, i.e., slightly lower compared to the use of turbulence promoter alone. The reason for this result can be found in the fact that flux increase is more affected by the presence of a Kenics static mixer while sparging results in an increase of energy used during microfiltration. By comparing combined method with turbulence promoter flux enhancement method ER is negative (-7%) which can be explained by increased power consumption for air flow with moderate contribution to the flux increase. Another confirmation for this fact can be found by comparing energy consumption values for combined method with energy consumption in the case of two-phase flow. In this instance energy reduction (ER) is 22% that demonstrates that turbulence promoter is more efficient compared to two phase flow. Antimicrobial activity of Bacillus velezensis biomass against phytopathogenic isolates Xanthomonas campestris was preserved under different fouling reduction methods.

Keywords: Bacillus velezensis, microfiltration, static mixer, two-phase flow

Procedia PDF Downloads 118

Authors: A. Falenski, A. Kaesbohrer, M. Filter

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Introduction: Risk assessments can be performed in various ways and in different degrees of complexity. In order to assess risks associated with imported foods additional information needs to be taken into account compared to a risk assessment on regional products. The present review is an overview on currently available best practise approaches and data sources used for food import risk assessments (IRAs). Methods: A literature review has been performed. PubMed was searched for articles about food IRAs published in the years 2004 to 2014 (English and German texts only, search string “(English [la] OR German [la]) (2004:2014 [dp]) import [ti] risk”). Titles and abstracts were screened for import risks in the context of IRAs. The finally selected publications were analysed according to a predefined questionnaire extracting the following information: risk assessment guidelines followed, modelling methods used, data and software applied, existence of an analysis of uncertainty and variability. IRAs cited in these publications were also included in the analysis. Results: The PubMed search resulted in 49 publications, 17 of which contained information about import risks and risk assessments. Within these 19 cross references were identified to be of interest for the present study. These included original articles, reviews and guidelines. At least one of the guidelines of the World Organisation for Animal Health (OIE) and the Codex Alimentarius Commission were referenced in any of the IRAs, either for import of animals or for imports concerning foods, respectively. Interestingly, also a combination of both was used to assess the risk associated with the import of live animals serving as the source of food. Methods ranged from full quantitative IRAs using probabilistic models and dose-response models to qualitative IRA in which decision trees or severity tables were set up using parameter estimations based on expert opinions. Calculations were done using @Risk, R or Excel. Most heterogeneous was the type of data used, ranging from general information on imported goods (food, live animals) to pathogen prevalence in the country of origin. These data were either publicly available in databases or lists (e.g., OIE WAHID and Handystatus II, FAOSTAT, Eurostat, TRACES), accessible on a national level (e.g., herd information) or only open to a small group of people (flight passenger import data at national airport customs office). In the IRAs, an uncertainty analysis has been mentioned in some cases, but calculations have been performed only in a few cases. Conclusion: The current state-of-the-art in the assessment of risks of imported foods is characterized by a great heterogeneity in relation to general methodology and data used. Often information is gathered on a case-by-case basis and reformatted by hand in order to perform the IRA. This analysis therefore illustrates the need for a flexible, modular framework supporting the connection of existing data sources with data analysis and modelling tools. Such an infrastructure could pave the way to IRA workflows applicable ad-hoc, e.g. in case of a crisis situation.

Keywords: import risk assessment, review, tools, food import

Procedia PDF Downloads 302

Authors: K. S. Al-Mugren, A. Yahya, S. Alodah, R. Alharbi, S. H. Almsaid , A. Alqahtani, H. Jaber, A. Basaqer, N. Alajra, N. Almoghati, A. Alsalman, Khalid Alharbi

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Context: The production of biofertilizers from naturally occurring microorganisms is an area of research that aims to enhance agricultural practices by utilizing local resources. This research project focuses on isolating and screening indigenous microorganisms with PK-fixing and phosphate solubilizing characteristics from local sources. Research Aim: The aim of this project is to develop a biofertilizer product using indigenous microorganisms and composted agro waste as a carrier. The objective is to enhance crop productivity and soil fertility through the application of biofertilizers. Methodology: The research methodology includes several key steps. Firstly, indigenous microorganisms will be isolated from local resources using the ten-fold serial dilutions technique. Screening assays will be conducted to identify microorganisms with phosphate solubilizing and PK-fixing activities. Agro-waste materials will be collected from local agricultural sources, and composting experiments will be conducted to convert them into organic matter-rich compost. Physicochemical analysis will be performed to assess the composition of the composted agro-waste. Gamma and X-ray irradiation will be used to sterilize the carrier material. The sterilized carrier will be tested for sterility using the ten-fold serial dilutions technique. Finally, selected indigenous microorganisms will be developed into biofertilizer products. Findings: The research aims to find suitable indigenous microorganisms with phosphate solubilizing and PK-fixing characteristics for biofertilizer production. Additionally, the research aims to assess the suitability of composted agro waste as a carrier for biofertilizers. The impact of gamma irradiation sterilization on pathogen elimination will also be investigated. Theoretical Importance: This research contributes to the understanding of utilizing indigenous microorganisms and composted agro waste for biofertilizer production. It expands knowledge on the potential benefits of biofertilizers in enhancing crop productivity and soil fertility. Data Collection and Analysis Procedures: The data collection process involves isolating indigenous microorganisms, conducting screening assays, collecting and composting agro waste, analyzing the physicochemical composition of composted agro waste, and testing carrier sterilization. The analysis procedures include assessing the abilities of indigenous microorganisms, evaluating the composition of composted agro waste, and determining the sterility of the carrier material. Conclusion: The research project aims to develop biofertilizer products using indigenous microorganisms and composted agro waste as a carrier. Through the isolation and screening of indigenous microorganisms, the project aims to enhance crop productivity and soil fertility by utilizing local resources. The research findings will contribute to the understanding of the suitability of composted agro waste as a carrier and the efficacy of gamma irradiation sterilization. The research outcomes will have theoretical importance in the field of biofertilizer production and agricultural practices.

Keywords: biofertilizer, microorganisms, agro waste, nuclear technologies

Procedia PDF Downloads 141

Authors: Anja I. Petrov, Milica B. Veljković, Marija M. Ćorović, Ana D. Milivojević, Milica B. Simović, Katarina M. Banjanac, Dejan I. Bezbradica

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One of the fundamental requirements for overall human well-being is a stable and balanced microbiome. Aside from the microorganisms that reside within the body, a large number of microorganisms, especially bacteria, swarming the human skin is in homeostasis with the host and represents a skin microbiota. Even though the immune system of the skin is capable of distinguishing between commensal and potentially harmful transient bacteria, the cutaneous microbial balance can be disrupted under certain circumstances. In that case, a reduction in the skin microbiota diversity, as well as changes in metabolic activity, results in dermal infections and inflammation. Probiotics and prebiotics have the potential to play a significant role in the treatment of these skin disorders. The most common resident bacteria found on the skin, Staphylococcus epidermidis, can act as a potential skin probiotic, contributing to the protection of healthy skin from pathogen colonization, such as Staphylococcus aureus, which is related to atopic dermatitis exacerbation. However, as it is difficult to meet regulations in cosmetic products, another therapy approach could be topical prebiotic supplementation of the skin microbiota. In recent research, polyphenols are attracting scientists' interest as biomolecules with possible prebiotic effects on the skin microbiota. This research aimed to determine how herbal extracts rich in different polyphenolic compounds (lemon balm, St. John's wort, coltsfoot, pine needle, and yarrow) affected the growth of S. epidermidis and S. aureus. The first part of the study involved screening plants to determine if they could be regarded as probable candidates to be skin prebiotics. The effect of each plant on bacterial growth was examined by supplementing the nutrient medium with their extracts and comparing it with control samples (without extract). The results obtained after 24 h of incubation showed that all tested extracts influenced the growth of the examined bacteria to some extent. Since lemon balm and St. John's wort extracts displayed bactericidal activity against S. epidermidis, whereas coltsfoot inhibited both bacteria equally, they were not explored further. On the other hand, pine needles and yarrow extract led to an increase in S. epidermidis/S. aureus ratio, making them prospective candidates to be used as skin prebiotics. By examining the prebiotic effect of two extracts at different concentrations, it was revealed that, in the case of yarrow, 0.1% of extract dry matter in the fermentation medium was optimal, while for the pine needle extract, a concentration of 0.05% was preferred, since it selectively stimulated S. epidermidis growth and inhibited S. aureus proliferation. Additionally, the total polyphenols and flavonoid content of the two extracts were determined, revealing different concentrations and polyphenol profiles. Since yarrow and pine extracts affected the growth of skin bacteria in a dose-dependent manner, by carefully selecting the quantities of these extracts, and thus polyphenols content, it is possible to achieve desirable alterations of skin microbiota composition, which may be suitable for the treatment of atopic dermatitis.

Keywords: herbal extracts, polyphenols, skin microbiota, skin prebiotics

Procedia PDF Downloads 175

Authors: Geisa Lima Mesquita Zambrosi, Maisa Ciampi Guillardi, Flávia Rodrigues Patrício, Oliveiro Guerreiro Filho

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Certified agricultural products are important components of the food industry. However, certifiers have been expanding the list of restricted or prohibited pesticides, limiting the options of products for phytosanitary control of plant diseases, but without offering alternatives to the farmers. Soybean and coffee leaf rust, brown eye spots, and Phoma leaf spots are the main fungal diseases that pose a serious threat to soybean and coffee cultivation worldwide. In conventional farming systems, these diseases are controlled by using synthetic fungicides, which, in addition to intensifying the occurrence of fungal resistance, are highly toxic to the environment, farmers, and consumers. In organic, agroecological, or regenerative farming systems, product options for plant protection are limited, being available only copper-based compounds, and biodefensivesornon-standard homemade products. Therefore, there is a growing demand for effective bioprotectors with low environmental impact for adoption in more sustainable agricultural systems. Then, to contribute to covering such a gap, we have developed a compound based on plant extracts and metallic elements for foliar application. This product has both biostimulant and bioprotective action, which promotes sustainable disease control, increases productivity as well as reduces damage to the environment. The product's components have complementary mechanisms that promote protection against the disease by directly acting on the pathogens and activating the plant's natural defense system. The protective ability of the product against three coffee diseases (coffee leaf rust, brown eye spot, and Phoma leaf spot) and against soybean rust disease was evaluated, in addition to its ability to promote plant growth. Our goal is to offer an effective alternative to control the main coffee fungal diseases and soybean fungal diseases, with a biostimulant effect and low toxicity. The proposed product can also be part of the integrated management of coffee and soybean diseases in conventional farming associated with chemical and biological pesticides, offering the market a sustainable coffee and soybean with high added value and low residue content. Experiments were carried out under controlled conditions to evaluate the effectiveness of the product in controlling rust, phoma, and cercosporiosis in comparison to control-inoculated plants that did not receive the product. The in vitro and in vivo effects of the product on the pathogen were evaluated using light microscopy and scanning electron microscopy, respectively. The fungistatic action of the product was demonstrated by a reduction of 85% and 95% in spore germination and disease symptoms severity on the leaves of coffee plants, respectively. The formulation had both a protective effect, acting to prevent infection by coffee leaf rust, and a curative effect, reducing the rust symptoms after its establishment.

Keywords: plant disease, natural fungicide, plant health, sustainability, alternative disease management

Procedia PDF Downloads 43

Authors: Z. Yousefniayejahr, S. Bolognini, A. Bonini, C. Campobasso, N. Poma, F. Vivaldi, M. Di Luca, A. Tavanti, F. Di Francesco

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Pathogenic bacteria represent a threat to healthcare systems and the food industry because their rapid detection remains challenging. Electrochemical biosensors are gaining prominence as a novel technology for the detection of pathogens due to intrinsic features such as low cost, rapid response time, and portability, which make them a valuable alternative to traditional methodologies. These sensors use biorecognition elements that are crucial for the identification of specific bacteria. In this context, bacteriophages are promising tools for their inherent high selectivity towards bacterial hosts, which is of fundamental importance when detecting bacterial pathogens in complex biological samples. In this study, we present the development of a low-cost and portable sensor based on the Zeno phage for the rapid detection of Staphylococcus aureus. Screen-printed gold electrodes functionalized with the Zeno phage were used, and electrochemical impedance spectroscopy was applied to evaluate the change of the charge transfer resistance (Rct) as a result of the interaction with S. aureus MRSA ATCC 43300. The phage-based biosensor showed a linear range from 101 to 104 CFU/mL with a 20-minute response time and a limit of detection (LOD) of 1.2 CFU/mL under physiological conditions. The biosensor’s ability to recognize various strains of staphylococci was also successfully demonstrated in the presence of clinical isolates collected from different geographic areas. Assays using S. epidermidis were also carried out to verify the species-specificity of the phage sensor. We only observed a remarkable change of the Rct in the presence of the target S. aureus bacteria, while no substantial binding to S. epidermidis occurred. This confirmed that the Zeno phage sensor only targets S. aureus species within the genus Staphylococcus. In addition, the biosensor's specificity with respect to other bacterial species, including gram-positive bacteria like Enterococcus faecium and the gram-negative bacterium Pseudomonas aeruginosa, was evaluated, and a non-significant impedimetric signal was observed. Notably, the biosensor successfully identified S. aureus bacterial cells in a complex matrix such as a nasal swab, opening the possibility of its use in a real-case scenario. We diluted different concentrations of S. aureus from 108 to 100 CFU/mL with a ratio of 1:10 in the nasal swap matrices collected from healthy donors. Three different sensors were applied to measure various concentrations of bacteria. Our sensor indicated high selectivity to detect S. aureus in biological matrices compared to time-consuming traditional methods, such as enzyme-linked immunosorbent assay (ELISA), polymerase chain reaction (PCR), and radioimmunoassay (RIA), etc. With the aim to study the possibility to use this biosensor to address the challenge associated to pathogen detection, ongoing research is focused on the assessment of the biosensor’s analytical performances in different biological samples and the discovery of new phage bioreceptors.

Keywords: electrochemical impedance spectroscopy, bacteriophage, biosensor, Staphylococcus aureus

Procedia PDF Downloads 67