Biomedical subjects

S D Pillai

Publications and source records attributed to S D Pillai.

At least 19 recordsLinked to original sources

PubMed2006

Cultural and Immunological Detection Methods for Salmonella spp. in Animal Feeds - A Review.

Food-borne salmonellosis continues to be a major public health concern, and contamination with Salmonella spp. in pre-harvest animal production is considered a primary contributor to this problem. Animal feeds can easily become contaminated during primary production, feed mixing and processing as well as during feeding. Consequently, monitoring and surveillance of feeds and feed ingredients for Salmonella spp. contamination may be useful or necessary in the prevention and control of this organism. Cultural and immunological detection methods for salmonellae have been used or suggested as possible approaches for use in animal feeds. Cultural methods remain advantageous owing to their ability to detect viable bacterial cells, while immunological methods have the capability of detecting nonculturable bacterial cells. Advancements and improvements in both methodologies offer opportunities for eventual routine use of these detection technologies in animal feed assays.

Animal Feed↗

PubMed2005

Polymerase chain reaction detection of foodborne Salmonella spp. in animal feeds.

Foodborne salmonellosis continues to be a public health issue of considerable concern. Animal feed has been a major link in pre-harvest food animal production. Although monitoring systems and control measures are available to limit Salmonella spp. contamination on animal feeds detection methodology is relatively time consuming in the context of time inputs for feed processing and mixing. Current cultural methods of Salmonella spp. detection in feeds require several days for confirmation. This amount of time represents significant problems if control measures are to be effectively implemented in a fashion that keeps feed processing costs low. Molecular methods offer improved sensitivity and potential reduction in assay time. In particular, several commercial polymerase chain reaction (PCR) assays, and combined PCR-hybridization assays have been suggested as possible means to implement more rapid detection of Salmonella spp. extracted from animal feeds. It has now become possible to rapidly detect and confirm the presence of foodborne Salmonella spp. in feed matrices by commercial amplification detection systems. The primary challenges remaining are to develop more reliable recovery and extraction procedures for routine processing of samples from a wide variety of feed matrices and apply molecular techniques for assessing physiological status of Salmonella spp. contaminants in animal feeds.

Animal Feed↗

PubMed2005

Concentrations of Escherichia coli and genetic diversity and antibiotic resistance profiling of Salmonella isolated from irrigation water, packing shed equipment, and fresh produce in Texas.

Fresh produce has been repeatedly implicated as a vehicle in the transmission of foodborne gastroenteritis. In an effort to assess the risk factors involved in the contamination of fresh produce with pathogenic bacteria, a total of 1,257 samples were collected from cantaloupe, oranges, and parsley (both in the field and after processing) and from the environment (i.e., irrigation water, soil, equipment, etc.). Samples were collected twice per season from two production farms per commodity and analyzed for the presence of Salmonella and Escherichia coli. E. coli was detected on all types of commodities (cantaloupe, oranges, and parsley), in irrigation water, and on equipment surfaces. A total of 25 Salmonella isolates were found: 16 from irrigation water, 6 from packing shed equipment, and 3 from washed cantaloupes. Salmonella was not detected on oranges or parsley. Serotyping, pulsed-field gel electrophoresis (PFGE), and repetitive element sequence-based PCR (rep-PCR) assays were applied to all Salmonella isolates to evaluate the genetic diversity of the isolates and to determine relationships between sources of contamination. Using PFGE, Salmonella isolates obtained from irrigation water and equipment were determined to be different from cantaloupe isolates; however, DNA fingerprinting did not conclusively define relationships between contamination sources. All Salmonella isolates were subjected to antimicrobial susceptibility testing using the disk diffusion method, and 20% (5 of 25) of the isolates had intermediate sensitivity to streptomycin. One Salmonella isolate from cantaloupe was resistant to streptomycin.

Anti-Bacterial Agents↗

PubMed2005

Survival of salmonella transformed to express green fluorescent protein on Italian parsley as affected by processing and storage.

To study the effect of processing and storage parameters on the survival of Salmonella on fresh Italian parsley, parsley bunches were dipped for 3 or 15 min in suspensions that were preequilibrated to 5, 25, or 35 degrees C and inoculated with Salmonella transformed to express enhanced green fluorescent protein. Loosely attached and/or associated, strongly attached and/or associated, and internalized and/or entrapped Salmonella cells were enumerated over 0, 1, and 7 days of storage at 25 degrees C and over 0, 1, 7, 14, and 30 days of storage at 4 degrees C using surface-plating procedures. Leaf sections obtained from samples after 0, 1, and 7 days of storage were examined using confocal scanning laser microscopy. Temperature of the dip suspension had little effect on the attachment and survival of Salmonella cells on parsley. Regardless of the temperature or duration of dip, Salmonella was internalized. Immersion for longer times resulted in higher numbers of attached and internalized cells. Microscopic observations supported these results and revealed Salmonella cells near the stomata and within cracks in the cuticle. Storage temperature had the greatest impact on the survival of Salmonella cells on parsley. When stored at 25 degrees C, parsley had a shelf life of 7 days, and Salmonella populations significantly increased over the 7 days of storage. For parsley stored at 4 degrees C, numbers of Salmonella cells decreased over days 0, 1, and 7. After 7 days of storage, there were no viable internalized Salmonella cells detected. Storage temperature represents an important control point for the safety of fresh parsley.

Bacterial Adhesion↗

PubMed2004

Development and evaluation of methods to detect coliphages in large volumes of water.

New and improved methods have been developed to detect somatic and male-specific coliphages in large volumes of water by single agar layer (SAL), enrichment and membrane filter methods. Somatic coliphages were detected efficiently on E. coli hosts C and CN13, male-specific coliphages were detected more efficiently on E. coli Famp than on Salmonella typhimurium WG49 and both types of coliphages were detected simultaneously on E. coli C3000. For water volumes of up to 100 ml, the SAL method was efficient and reliable. For water volumes of <1 L and as many as 10 multiple 1 L volumes, the enrichment method was efficient in detecting very low numbers of coliphages. Membrane filter methods, in which coliphages were adsorbed to and eluted from filters, also were relatively efficient, but they were less efficient than SAL and enrichment methods and were considered to be more cumbersome. For filter adsorption-elution methods, coliphage recoveries were most efficient for cellulose ester filters, less efficient for electropositive 1 MDS filters and least efficient for a direct membrane filter method. Overall, the enrichment method was preferred because of its ability to easily and rapidly detect low levels of coliphages in large sample volumes by either presence-absence or most probable number quantification.

Cellulose↗

PubMed2003

Use of a Salmonella typhimurium hilA fusion strain to assess effects of environmental fresh water sources on virulence gene expression.

Many fruits and vegetables are irrigated with water from rivers, lakes and even wastewater systems. Irrigation may be a route for the introduction of Salmonella. Our objectives in this study were to determine survivability and virulence expression in a strain of Salmonella typhimurium when exposed to environmental water sources. Virulence expression was measured using a beta-galactosidase assay on a hilA:lacZY fusion strain of S. typhimurium. Water samples for environmental impact studies were taken from a local pond and specific sites along the Rio Grande River, which serves as a source of irrigation water in southern Texas. There was a significant difference (p<0.05) of virulence expression among the water sites. Certain regions along the Rio Grande River yielded greater amounts of beta-galactosidase activity than others. All sites yielded at least a two-fold greater virulence response than S. typhimurium grown in brain heart infusion. Salmonella survivors were enumerated as colony forming units (CFU)/ml as plated on a selective medium for the duration of 1 week and beta-galactosidase assays were performed to determine a possible correlation between culturable cells and virulence gene expression. Bacterial cells remained viable but decreased after 7 days incubation. In conclusion, water sampled at specific locations and at different times water samples exhibited differences in virulence expression in S. typhimurium.

Agriculture↗

PubMed2003

Monitoring and identifying antibiotic resistance mechanisms in bacteria.

Sub-therapeutic administration of antibiotics to animals is under intense scrutiny because they contribute to the dissemination of antibiotic-resistant bacteria into the food chain. Studies suggest that there is a link between the agricultural use of antibiotics and antibiotic-resistant human infections. Antibiotic-resistant organisms from animal and human wastes reenter the human and animal populations through a number of pathways including natural waters, irrigation water, drinking water, and vegetables and foods. Antibiotic usage in the United States for animal production (disease prevention and growth promotion) is estimated to be 18 million pounds annually. As much as 25 to 75% of the antibiotics administered to feedlot animals are excreted unaltered in feces. Because about 180 million dry tons of livestock and poultry waste is generated annually in the United States, it is not surprising that animal-derived antibiotic-resistant organisms are found contaminating groundwater, surface water, and food crops. It is extremely important to clearly understand the molecular mechanisms that could potentially cause lateral or horizontal gene transfer of antibiotic resistance genes among bacteria. Once the mechanisms and magnitude of resistance gene transfer are clearly understood and quantified, strategies can be instituted to reduce the potential for dissemination of these genes.

Animal Husbandry↗

PubMed2002

Comparison of media and a liquid detergent extraction step on bacterial recovery from animal feeds.

The objective of this study was to evaluate the recovery of Tryptic soy agar (TSA) or low nutrient R2 bacterial populations from fresh feeds and animal feeds held in long-term cold storage after extraction with a detergent-based solution combined with repeated centrifugation. Repeated centrifugation after exposure to a detergent-based solution resulted in recoveries generally no more than 10% of the microbial population in the original feed. Feed R2A populations were significantly correlated (P<0.01) with TSA populations enumerated from the feed and feed extracts, as well as R2A populations from feed extracts. Similarly, TSA bacterial populations from feed were significantly correlated with TSA populations from extracts.

Animal Feed↗

PubMed2001

Presence of bacteriophages in animal feed as indicators of fecal contamination.

The objectives of this study were to determine if indigenous male specific and somatic bacteriophages could be detected in animal feeds and if isolated phages contained RNA or DNA. Seven fresh feeds, 2 fresh feed ingredients, 7 stored feeds, 2 stored feed ingredients, and 8 samples of poultry diets suspected to contain Salmonella spp. were enriched and spot plated for indigenous phages using Escherichia coli Famp and CN-13 as hosts. Bacteriophage numbers were below detection without enrichment, but both male specific and somatic coliphages were detected in all animal feeds, feed ingredients, and poultry diets after 16 h of enrichment, even after the samples had been stored for 14 months of storage at -20 C. Five out of 9 fresh feeds and 7 out of 8 stored feeds contained RNA somatic phages.

Animal Feed↗

PubMed2000

Failure to identify non-bovine reservoirs of Mycobacterium bovis in a region with a history of infected dairy-cattle herds.

The State of Texas had the most (cumulative) tuberculous cattle herds of any state in the United States during the decade ending in 1997. Of the cumulative 18 infected herds in Texas, 12 herds were concentrated in El Paso County (designated the 'El Paso milkshed'). To identify whether non-bovine reservoirs were a source of Mycobacterium bovis infection of cattle in this region, an investigation was conducted on the premises of 14 dairy herds (12 tuberculous and 2 non-affected herds) between May 1995 and June 1997. None of the 670 mammalian, avian and environmental (soil, water and air) samples collected and cultured from the premises of these herds was positive for the presence of M. bovis. None of the 119 human urine samples obtained from employees of these dairies was culture positive for M. bovis. Of 124 dairy-farm workers with tuberculin skin-test results, 48 showed positive test results. There was, however, no difference in percentages of positive skin-test results between farms without, and farms having, bovine tuberculosis within the last two years or longer. The percentage of positive reactions did not increase with length of time employed at a dairy with a history of confirmed tuberculosis. These findings suggest that non-bovine reservoirs appear not to be a factor responsible for tuberculosis of cattle in the El Paso milkshed.

Adult↗

PubMed2000

Efficacy of a commercial polymerase chain reaction-based assay for detection of Salmonella spp. in animal feeds.

Salmonellosis is a cyclic problem in the food industry, to which animal feed has been contributory. Current conventional methods of Salmonella spp. detection require 96 h for detection and confirmation. With modern and just-in-time production schedules, a 96-h hold represents a significant expense in storage and decontamination. The commercially available assay, 'BAX for Screening/Salmonella' (BAX), is based on the principle of the polymerase chain reaction and may represent a significant decrease in assay time. Seven fresh feed formulations, two fresh feed ingredients, seven stored feeds and two stored feed ingredients were artificially contaminated with a primary poultry isolate of Salmonella typhimurium and analysed by conventional and BAX methodology. The results of BAX agreed with conventional plating results for 16 of 18 samples spiked with 1200 cfu 10 g(-1) of feed and 13 of 18 samples spiked with 40 cfu 10 g(-1) of feed. Indigenous Salmonella spp. were detected in five of eight samples of poultry diets by conventional methods. With BAX, Salmonella spp. could not be detected in any of the samples after only 7 h of enrichment but could be detected in two dietary samples after 13 h of enrichment and four dietary samples after 24 h of enrichment. Specific sequences of salmonella DNA that were extracted from poultry diets could be detected with BAX.

Animal Feed↗

PubMed2000

Recovery of a marker strain of Salmonella typhimurium in litter and aerosols from isolation rooms containing infected chickens.

Screening of poultry flocks for foodborne pathogen Salmonella contamination is critical for Salmonella control in preharvest stages of poultry production. In this study, two sampling methods (litter and air filter) were compared for detection of S. typhimurium from experimentally infected chicks some of which had received either a probiotic competitive exclusion culture or transfer of cecal contents from salmonellae-free adult birds. At 4, 9, and 11 days after inoculation, S. typhimurium samples were enumerated by selective plating. For both types of sampling, the control birds yielded the greatest levels of environmental contamination followed by the samples from the probiotic inoculated birds with the birds receiving the cecal transfer culture having the lowest levels of contamination. Although the two sampling methods responded in a similar fashion, detection sensitivity needs to be increased for air filter sampling.

Aerosols↗

PubMed1999

Conventional and molecular methods for understanding probiotic bacteria functionality in gastrointestinal tracts.

The recent successes of probiotic application to limit colonization of foodborne pathogens in the gastrointestinal tracts of food animals ensures continued commercialization and widespread use of such cultures. Given that the the fermentation response and ecological balance of the probiotic consortium appears to be essential for the effectiveness of the cultures, it is essential to develop a methodology to accurately identify and quantitate these organisms during commercial production as well as successful in vivo colonization after administration. However, if further optimization of the effectiveness of defined cultures is to be achieved, methods to assess expression of key metabolic processes occurring during establishment of the probiotic culture as well as its subsequent ability to limit foodborne pathogen colonization are needed. Conventional methods to study individual probiotic gastrointestinal organisms include selective plating to identify specific nutritional groups, but the requirement of strict anaerobiosis for the obligate anaerobic members of these cultures can confound sample handling and preparation. Immunological methods can circumvent some of these problems but are somewhat limited for assessing functionality. The main advantage of using molecular tools is that the genetic diversity of the microflora, as well as their gene activity data are obtainable, both at the community level and at the single species level. Methods are currently available that permit studying individual members of microbial consortia, fluxes in community diversity, spatial distribution of consortia members, and the expression of specific microbial genes within communities. These methods involve the utilization of both DNA- and RNA-targeted probes, gene amplification protocols, and mRNA analysis. The study of mechanisms and functionality can only enhance the potential of probiotic cultures for limiting foodborne pathogen colonization.

Anaerobiosis↗

PubMed1998

Delineating the specific influence of virus isoelectric point and size on virus adsorption and transport through sandy soils.

Many of the factors controlling viral transport and survival within the subsurface are still poorly understood. In order to identify the precise influence of viral isoelectric point on viral adsorption onto aquifer sediment material, we employed five different spherical bacteriophages (MS2, PRD1, Q beta, phi X174, and PM2) having differing isoelectric points (pI 3.9, 4.2, 5.3, 6.6, and 7.3 respectively) in laboratory viral transport studies. We employed conventional batch flowthrough columns, as well as a novel continuously recirculating column, in these studies. In a 0.78-m batch flowthrough column, the smaller phages (MS2, phi X174, and Q beta), which had similar diameters, exhibited maximum effluent concentration/initial concentration values that correlated exactly with their isoelectric points. In the continuously recirculating column, viral adsorption was negatively correlated with the isoelectric points of the viruses. A model of virus migration in the soil columns was created by using a one-dimensional transport model in which kinetic sorption was used. The data suggest that the isoelectric point of a virus is the predetermining factor controlling viral adsorption within aquifers. The data also suggest that when virus particles are more than 60 nm in diameter, viral dimensions become the overriding factor.

Adsorption↗

PubMed1997

Rapid molecular detection of microbial pathogens: breakthroughs and challenges.

Microbiological contamination of foods and drinking water is a global problem, and a significant amount of expense is being incurred as a result of such contamination. The microorganisms associated with almost half of all disease outbreaks still go unidentified, primarily as a result of inadequate monitoring and surveillance. Though significant improvements have been made in refining molecular methods for detecting infectious agents, a majority of these methods are being employed only on clinical samples where pathogen densities are much higher than those found in environmental and food samples. Comparative evaluations of the various protocols in terms of cost, sensitivity, specificity, speed, and reproducibility need to be undertaken so that the true applicability of these methods can determined. In the future, molecular methods, especially gene amplifications and in situ hybridizations, will find increasing applications in the differentiation of viable and non-viable organisms, in predicting antimicrobial resistance, and in the identification and characterization of unculturable microorganisms. Though molecular detection methods will not totally replace conventional methods, they will significantly enhance our ability to detect microbial pathogens rapidly.

Animals↗

PubMed1997

A rapid viability assay for Cryptosporidium oocysts and Giardia cysts for use in conjunction with indirect fluorescent antibody detection.

There is an urgent need to develop rapid methods to determine the viability of Cryptosporidium oocysts and Giardia cysts in environmental samples, especially water. The inclusion of the vital dye propidium iodide (PI) by oocysts and cysts has been previously shown to correlate well with nonviability. The ability of nonviable oocysts and cysts to include PI has been employed to develop a rapid viability determination method that could be used in conjunction with the current indirect fluorescent antibody (IFA) method for detecting oocysts and cysts. The efficacy of this PI-IFA method to detect and determine the viability status of oocysts/cysts has been tested using oocyst samples inactivated by three different approaches. The ability to incorporate PI staining with IFA detection provides the advantage of both detection and viability determination at the same time, using the same sample.

Animals↗

PubMed1996

Occurrence of airborne bacteria and pathogen indicators during land application of sewage sludge.

Glass impingers (AGI-30) were used at a commercial sludge application site to determine the levels of airborne bacteria and pathogen indicators. Even though heterotrophic bacteria averaged 10(5) CFU/m3, none of the sites showed the presence of Salmonella spp. or indicators such as fecal coliforms or coliphages. Indicators such as H2S producers and pathogenic clostridia were present in locations having significant physical agitation of the sludge material. PCR-based ribotyping using the 16S-23S interspacer region is a promising method to identify the genetic relatedness and origins of airborne clostridia.

Air Microbiology↗

PubMed1995

Strategies to accelerate the applicability of gene amplification protocols for pathogen detection in meat and meat products.

Traditionally, microbiological testing of meat products has involved isolating microorganisms and performing specific biochemical, and in some cases serological, tests to confirm the presence or absence of suspected food-borne pathogens. Given the public attention meat products have received as sources of food-borne disease, there has been considerable interest in the application of rapid detection techniques that require hours rather than days for completion. Theoretically, rapid detection methods could reduce the time from the initial sampling to confirmation so that conclusive results would be available by the time to process the meat product. Both direct gene probe hybridization as well as gene amplification methods show promise as rapid detection techniques. At present, direct gene probe hybridization are being commercially utilized to confirm the presence of a suspected pathogen. A number of gene amplification protocols for detecting food-borne bacterial pathogens have been published. However, many of these studies have utilized spiked samples rather than naturally contaminated samples and many of them have involved extended template extraction/purification methodologies. There is still only a very limited amount of information on the efficacies of the various protocols in detecting bacterial pathogens, especially toxigenic Escherichia coli, Salmonella spp., Campylobacter spp., and Listeria spp., in naturally contaminated food samples. In order to develop gene amplification protocols that have relevance to the meat industry, there must be a concerted effort to utilize naturally contaminated samples in the development and evaluation of protocols, as well as to initiate multilaboratory round robin evaluations of select protocols. Availability of multilaboratory tested methodologies would provide a means to design pathogen detection strategies at the quality control level rather than an end product confirmatory response to an already documented outbreak.

Animals↗