Incompatibility (Inc) HI2 plasmids are large (typically > 200 kb), transmissible plasmids that encode antimicrobial resistance (AMR), heavy metal resistance (HMR) and disinfectants/biocide resistance (DBR). To better understand the distribution and diversity of resistance-encoding genes among IncHI2 plasmids, computational approaches were used to evaluate resistance and transfer-associated genes among the plasmids. Complete IncHI2 plasmid (N = 667) sequences were extracted from GenBank and analyzed using AMRFinderPlus, IntegronFinder and Plasmid Transfer Factor database. The most common IncHI2-carrying genera included Enterobacter (N = 209), Escherichia (N = 208), and Salmonella (N = 204). Resistance genes distribution was diverse, with plasmids from Escherichia and Salmonella showing general similarity in comparison to Enterobacter and other taxa, which grouped together. Plasmids from Enterobacter and other taxa had a higher prevalence of multiple mercury resistance genes and arsenic resistance gene, arsC, compared to Escherichia and Salmonella. For sulfonamide resistance, sul1 was more common among Enterobacter and other taxa, compared to sul2 and sul3 for Escherichia and Salmonella. Similar gene diversity trends were also observed for tetracyclines, quinolones, β-lactams, and colistin. Over 99% of plasmids carried at least 25 IncHI2-associated conjugal transfer genes. These findings highlight the diversity and dissemination potential for resistance across different enteric bacteria and value of computational-based approaches for the resistance-gene assessment.

Salmonella is estimated to cause over a million infections and ~400 deaths annually in the U.S. Salmonella enterica serotype Javiana strains (n = 409) that predominantly originated from the State of Arkansas over a six-year period (2003 to 2008) were studied. This period coincided with a rapid rise in the incidence of S. Javiana infections in the U.S. Children under the age of 10 displayed the highest prevalence of S. Javiana infections, regardless of sex or year of detection. Antimicrobial susceptibility to 15 different antimicrobials was assessed and 92% (n = 375) were resistant to at least one of the antimicrobials. Approximately 89% of the isolates were resistant to sulfisoxazole alone and 3% (n = 11) were resistant to different antimicrobials, including gentamicin, ciprofloxacin or ceftiofur. The pulsed-field gel electrophoresis (PFGE) analyses assessed the genotypic diversity and distribution of S. Javiana strains using XbaI restriction. Nine major clusters were identified and isolates from each group were digested with the restriction enzyme AvrII. Isolates with identical profiles of XbaI and AvrII were found to be disseminated in human populations. These distinct “types” of S. Javiana were persistent in human populations for multiple years. A subset of isolates (n = 19) with unique resistance phenotypes underwent plasmid and incompatibility (Inc) type analyses and the isolates resistant to more than one antimicrobial harbored multiple plasmids (<3 to 165 kb). Furthermore, these strains possessed 14 virulence genes, including pagC, cdtB, and iroN. The whole genome sequences (WGS) of 18 isolates that mostly originated from Arkansas from 2003 to 2011 were compared with isolates collected from different areas in the U.S. in 1999, indicating the perseverance of S. Javiana in disseminating antimicrobial resistance and virulence genes.

The quality of fecal specimens is one of the factors responsible for successful Clostridioides difficile infection (CDI) diagnosis. The quality depends largely on the storage conditions, including the temperature and time period. In this study, we organized the outputs of previous studies, filled experimental gaps in the knowledge of storage conditions, and introduced a pragmatic strategy for fecal storage for CDI diagnosis. A 5-step pathway was adopted to develop the fecal specimen storage strategy as follows: step 1, bibliomic analysis; step 2, experimental gap-filling; step 3, comparative evaluation; step 4, strategy development; step 5, internal review. Step 1 identified eight articles providing experimental information on the effects of fecal specimen storage conditions on the effectiveness of C. difficile detection methods. Step 2 provided additional quantitative data on C. difficile vegetative and spore cell viability and DNA stability. All previous and current results were compared (step 3). In step 4, fir general and nine special strategies were developed, followed by an internal review of the overall approaches (step 5). It is recommended to separate fecal samples into aliquots before testing and storing them. It is particularly recommended that fecal specimen samples be stored for CDI diagnosis at 4 °C for up to 60 days for all test methods.

Sixty-six Salmonella enterica serovar Typhimurium isolates carrying incompatibility group FIB (IncFIB) plasmids were sequenced to further characterize the IncFIB plasmid-encoded factors associated with virulence and antimicrobial resistance genes. In addition to the IncFIB plasmid, many of these isolates harbored additional plasmids encoding virulence and antimicrobial resistance genes.

Salmonella enterica is one of the most common bacterial foodborne pathogens in the United States, causing illnesses that range from self-limiting gastroenteritis to more severe, life threatening invasive disease. Many Salmonella strains contain plasmids that carry virulence, antimicrobial resistance, and/or transfer genes which allow them to adapt to diverse environments, and these can include incompatibility group (Inc) FIB plasmids. This study was undertaken to evaluate the genomic and phenotypic characteristics of IncFIB-positive Salmonella enterica serovar Typhimurium isolates from food animal sources, to identify their plasmid content, assess antimicrobial resistance and virulence properties, and compare their genotypic isolates with more recently isolated S. Typhimurium isolates from food animal sources. Methods: We identified 71 S. Typhimurium isolates that carried IncFIB plasmids. These isolates were subjected to whole genome sequencing and evaluated for bacteriocin production, antimicrobial susceptibility, the ability to transfer resistance plasmids, and a subset was evaluated for their ability to invade and persist in intestinal human epithelial cells. Results: Approximately 30% of isolates (n = 21) displayed bacteriocin inhibition of Escherichia coli strain J53. Bioinformatic analyses using PlasmidFinder software confirmed that all isolates contained IncFIB plasmids along with multiple other plasmid replicon types. Comparative analyses showed that all strains carried multiple antimicrobial resistance genes and virulence factors including iron acquisition genes, such as iucABCD (75%), iutA (94%), sitABCD (76%) and sitAB (100%). In 17 cases (71%), IncFIB plasmids, along with other plasmid replicon types, were able to conjugally transfer antimicrobial resistance and virulence genes to the susceptible recipient strain. For ten strains, persistence cell counts (27%) were noted to be significantly higher than invasion bacterial cell counts. When the genome sequences of the study isolates collected from 1998–2003 were compared to those published from subsequent years (2005–2018), overlapping genotypes were found, indicating the perseverance of IncFIB positive strains in food animal populations. This study confirms that IncFIB plasmids can play a potential role in disseminating antimicrobial resistance and virulence genes amongst bacteria from several food animal species.

Having reliable methods for detecting Shiga toxin-producing Escherichia coli (STEC) in foods is an important food safety goal. The majority of STEC outbreaks have involved either the O157:H7 serotype or one of six non-O157 serogroups, O26, O45, O103, O111, O121, and O145, termed "The Big Six." We have compared detection by PCR of the Shiga toxin genes stx1a and stx2a from STEC bacteria isolated from unclarified apple juice by simple centrifugation with the use of an immunocapture technique to minimize contaminants (such as pectin and polyphenols that may copurify with DNA) that may interfere with DNA amplification efficiencies and limit sensitivity. An internal control for successful immunocapture, DNA extraction, and PCR amplification was generated by introducing the pmRaspberry plasmid into an stx null strain, yielding an E. coli O45 pmRaspberry derivative that can be added to food samples directly. Using serial dilutions of a representative Big Six STEC in apple juice, our immunocapture method resulted in a 50% probability of detection value of 3.34, 2.25, and 4.25 CFU for detection by multiplex real-time PCR, growth on solid agar, and multiplex endpoint PCR, respectively. The time to result was 6.5 h, 9.5 h, and 1.5 days for immunocapture of Big Six STECs and detection by multiplex real-time PCR, endpoint PCR, and growth on solid agar, respectively. A set of 52 Big Six STEC isolates and 30 non-Big Six STEC strains was used to establish the inclusivity and exclusivity of the method. Finally, the ability to detect Big Six STEC contamination reliably was confirmed at 4.5 and 45 CFU/25-mL portions of refrigerated apple juice.

BACKGROUND:Antimicrobial resistance in Salmonella enterica is often plasmid encoded. A key resistance plasmid group is the incompatibility group (Inc) A/C plasmids that often carry multiple resistance determinants. Previous studies showed that IncA/C plasmids were often co-located with other plasmids. The current study was undertaken to evaluate the impact of plasmid co-carriage on antimicrobial resistance and plasmid transfer.METHODS:A total of 1267 Salmonella isolates, representing multiple serotypes and sources were previously subjected to susceptibility testing and 251 isolates with resistance to at least 5 antimicrobial agents were identified for further study. Each isolate was subjected to PCR-based replicon typing, and those with IncA/C plasmids were selected for plasmid isolation, PCR-based mapping of IncA/C plasmid backbone genes, and conjugation assays to evaluate resistance plasmid transferability.RESULTS:Of the 87 identified IncA/C positive isolates, approximately 75% carried a plasmid with another identified replicon type, with the most common being I1 (39%), FIA, FIIA, FIB and HI2 (each 15%). PCR-based mapping indicated significant diversity in IncA/C backbone content, especially in regions encoding transfer-associated and hypothetical proteins. Conjugation experiments showed that nearly 68% of the isolates transferred resistance plasmids, with 90% containing additional identified plasmids or larger (>50 kb) non-typeable plasmids.CONCLUSIONS:The majority of IncA/C-positive strains were able to conjugally transfer antimicrobial resistance to the recipient, encoded by IncA/C and/or co-carried plasmids. These findings highlight the importance of co-located plasmids for resistance dissemination either by directly transferring resistance genes or by potentially providing the needed conjugation machinery for IncA/C plasmid transfer.

ABSTRACT The draft genome sequences of eight Salmonella enterica isolates from various sources were evaluated for the influence of incompatibility group I1 (IncI1) plasmids on virulence. Strains SE142, SE143, SE144, and SE146 originated from swine, SE36N and SE89N from poultry-related sources, and SE991 and SE1148 from human patients.

Salmonella is a predominant foodborne pathogen in the United States and other countries. Mobile genetic elements such as plasmids allow Salmonella to adapt to external stress factors such as nutrient deprivation and host factors. Incompatibility group I1 (IncI1) plasmid-carrying Salmonella enterica strains were examined to determine the presence of plasmid-associated genes and their influence on phenotypic characteristics. The objective of this study was to understand the genetic determinants on IncI1 plasmids and their impact on antimicrobial susceptibility, competitive growth inhibition of Escherichia coli, and plasmid transfer. Primers were designed for genes that play a role in virulence, antimicrobial resistance, and plasmid transfer based on previously sequenced IncI1 plasmids. Polymerase chain reaction assays were conducted on 92 incompatibility group I1 (IncI1)-positive S. enterica strains. Phenotypic characterization included conjugation assays, antimicrobial susceptibility testing, and bacteriocin production based on the inhibition of growth of colicin-negative E. coli J53. The antimicrobial resistance genes aadA1, tetA, sul1, and blaCMY were detected in 88%, 87%, 80%, and 48% of the strains, respectively. Over half of the strains were resistant or intermediately resistant to streptomycin (85%), sulfonamides (76%), tetracycline (74%), and ampicillin (68%) and 57% of the strains inhibited growth of E. coli J53 strain. Among putative virulence genes, colicin-associated colI and cib were detected in 23% and 35% of strains and imm and ccdA were present in 58% and 54% of strains, respectively. Approximately 61% of strains contained plasmids that conjugally transferred antimicrobial resistance, including 83% where the recipient received IncI1 plasmids. Most of the strains carried an assortment of transfer associated (pil and tra) genes with between 63% and 99% of strains being positive for individual genes. Taken together the study affirms that IncI1 plasmids likely play roles in the dissemination of antimicrobial resistance and virulence-associated factors among enteric organisms.

Delayed sowing of wheat tends to reduce germination count and number of tillers/unit area because of sharp rise in temperature during tillering phase of the crop and ultimately decrease in yield (Soomro and Oad 2002). Hence, an attempt was made to cultivate weed free late sown wheat by use of fenoxaprop-p-ethyl at different doses in lieu of higher productivity and profitability from rice fallow areas.

Uttar Pradesh is the second largest producer accounting for about 16 per cent of total production. The area, production and productivity of pulses in UP including district Azamgarh is quite low as compared to other states in respect of national acreage and production. Among various constraints, poor crop management and protection technologies assume primary position. Considering the facts of low yield of pulses due to technological gap and various other constraints, Krishi Vigyan Kendra, Azamgarh of Uttar Pradesh conducted front line demonstration consequently five years on improved agricultural technologies of pulses i.e. pigeonpea, chickpea, field pea and lentil in scientific manner at farmers’ field during 2008-09 to 2012-13. A total of 241 demonstrations of pulses were conducted in an area of 90.0 hectares. The results of five years under front line demonstration on pulses revealed that the average grain yield of pigeonpea (19.1 q/ha), chickpea (19.1 q/ha) field pea (23.5 q/ha) and lentil (17.8 q/ ha) with their 56.5, 43.6, 40.8 and 45.1 per cent increase in yield over farmers practice were recorded under demonstration plots. Implementations of improved technological interventions in all demonstrated crops were also found remunerative in terms of B: C ratio over existing practices. The enhanced yield achieved through adoption of improved production and protection technology in pulses maintain the soil health, incremental sustainable development in production, enhancing nutritional securities and improves the livelihood of the farmers. The outcome of the trial inspired the farming communities to replace their old nondescriptive varieties with resistant and high yielding varieties and other production and protection related technological options which are being cultivated.

Standard methods to detect Escherichia coli contamination in food use the polymerase chain reaction (PCR) and agar culture plates. These methods require multiple incubation steps and take a long time to results. An improved rapid flow-cytometry based detection method was developed, using a fluorescence-labeled oligonucleotide probe specifically binding a16S rRNA sequence. The method positively detected 51 E. coli isolates as well as 4 Shigella species. All 27 non-E. coli strains tested gave negative results. Comparison of the new genetic assay with a total plate count (TPC) assay and agar plate counting indicated similar sensitivity, agreement between cytometry cell and colony counts. This method can detect a small number of E. coli cells in the presence of large numbers of other bacteria. This method can be used for rapid, economical, and stable detection of E. coli and Shigella contamination in the food industry and other contexts.

This study evaluated antimicrobial resistance and virulence factors in Salmonella enterica isolated from a turkey flock in which the birds were raised in an environment where antimicrobials were not administered to the birds, either through feed or water. Salmonella was isolated from turkeys and various environmental samples in the facility using conventional microbiological procedures. Isolates were serotyped and analyzed phenotypically by antimicrobial resistance profiling and genotypically by pulsed-field gel electrophoresis (PFGE) fingerprinting, integron analysis, plasmid profiling, replicon-based incompatibility (Inc) group typing, and virulence gene profiling. Ninety-five S. enterica isolates were isolated from cecal contents (n = 29), feed (n = 22), leftover feed (n = 13), litter (n = 12), drinkers (n = 10), environment (n = 8), and an insect. The following serotypes were identified: Montevideo (24%), Anatum (22%), Agona (17%), Kentucky and Worthington (12%), Senftenberg (11%), and rough phenotypes (3%). The majority of isolates (61/95; 64%) were susceptible to 12 antimicrobials tested; however, despite the absence of antimicrobials in the facility, approximately 36% of the isolates were resistant to two to five antimicrobials. Class 1 integrons were detected in 8% of the isolates. The integron sequence analysis revealed dihydrofolate reductase (dhfr) and aminoglycoside adenylyl transferase (aadA2) genes, which encode trimethoprim and streptomycin resistance, respectively. Furthermore, 71% of the isolates had at least one plasmid. There were five plasmid replicon types identified among the isolates, including IncI1, IncHI2, IncFIIA, IncB/O, and IncP, with variable prevalence among the serotypes. All 95 isolates tested polymerase chain reaction-positive for 19 virulence genes and negative for virD4 and virB4. The virulence gene profiles were similar within the isolates from the same serotype. Within particular serotypes, PFGE patterns revealed 100% similarity, even when the bacterial strains were isolated from different sources, indicating cross-colonization of sources within the turkey facility. On this antibiotic-free turkey farm, turkeys and feed appeared to be the major reservoirs of multidrug-resistant Salmonella, which harbored multiple virulence genes.

A field experiment was conducted during Rabi season of 2010-11 at Faizabad to assess the bio-efficacy of fenoxaprop and clodinofop on weeds and productivity of late sown wheat. Fenoxaprop at higher doses effectively suppressed weeds and produced significantly lowest weed dry weight but it did not result in enhancing nitrogen removal by the crop at lower doses. Post-emergence application of Whip Super (fenoxaprop) 120 g, fenoxaprop 100 and 120 g and clodinofop 60 g/ha in late sown wheat eliminated the crop weed competition from early crop growth stages which ultimately provided congenial condition to crop for better harnessing of available crop growth resources and gave profitable grain yields.

Clostridium perfringens is an important pathogen, causing food poisoning and other mild to severe infections in humans and animals. Some strains of C. perfringens contain conjugative plasmids, which may carry antimicrobial resistance and toxin genes. We studied genomic and plasmid diversity of 145 C. perfringens type A strains isolated from soils, foods, chickens, clinical samples, and domestic animals (porcine, bovine and canine), from different geographic areas in the United States between 1994 and 2006, using multiple-locus variable-number tandem repeat analysis (MLVA) and/or pulsed-field gel electrophoresis (PFGE). MLVA detected the genetic diversity in a majority of the isolates. PFGE, using SmaI and KspI, confirmed the MLVA results but also detected differences among the strains that could not be differentiated by MLVA. All of the PFGE profiles of the strains were different, except for a few of the epidemiologically related strains, which were identical. The PFGE profiles of strains isolated from the same domestic animal species were clustered more closely with each other than with other strains. However, a variety of C. perfringens strains with distinct genetic backgrounds were found among the clinical isolates. Variation was also observed in the size and number of plasmids in the strains. Primers for the internal fragment of a conjugative tcpH gene of C. perfringens plasmid pCPF4969 amplified identical size fragments from a majority of strains tested; and this gene hybridized to the various-sized plasmids of these strains. The sequences of the PCR-amplified tcpH genes from 12 strains showed diversity among the tcpH genes. Regardless of the sources of the isolates, the genetic diversity of C. perfringens extended to the plasmids carrying conjugative genes.

The area, production and productivity of pulses in UP including district Azamgarh is quite low as compared to other states in respect of national acreage and production. Among various constraints poor crop management and protection technologies assume primary position. Considering the facts of low yield of pulses due to technological gap and various other constraints Krishi Vigyan Kendra, Azamgarh conducted frontline demonstration continuously for five years on improved agricultural technologies of pulses viz pigeon pea, chickpea, field pea and lentil in scientific manner at farmers’ fields during 2008-09 to 2012-13. A total of 241 demonstrations of pulses were conducted in an area of 90.0 hectares. The results of five years under frontline demonstration on pulses revealed that the average grain yield of pigeon pea (19.1 q/ha), chickpea (19.7 q/ha) field pea (23.5 q/ha) and lentil (17.8 q/ha) with 56.5, 43.6, 40.8 and 45.1 per cent increase in yield respectively over farmers practice was recorded under demonstration plots. Implementation of improved technological interventions in all demonstrated crops were also found remunerative in terms of B:C ratio over existing practices. The enhanced yield achieved through adoption of improved production and protection technology in pulses maintained the soil health, incremental sustainable development in production, enhanced nutritional security and improved the livelihood of the farmers. The outcome of the trial inspired the farming community to replace their old non-descriptive varieties with resistant and high yielding varieties and other production and protection related technological options which were being implemented.

Field experiment was conducted during the winter season of 2009-10 at Agronomy Research Farm of ND University of Agriculture and Technology, Kumarganj, Faizabad, UP to study the effect of sequential application of different herbicides on late sown wheat of eastern Uttar Pradesh. Sequential application of pinoxaden 40 g followed by carfentrazone 25 g/ha (NH2SO4 1%) proved significantly effective in reducing density and biomass of weeds and gave complete control of grassy and broadleaf weeds without any phototoxic effect on crop as compared to carfentrazone at 35 days after sowing. The maximum grain yield 43.02 q/ha was recorded with completely weed free situation and it was statistically comparable with yield (41.6 q/ha) obtained under sequential application of pinoxaden 40 g followed by carfentrazone 25 g/ha (NH2SO4 1%) while lowest yield 31.4 q/ha in weedy condition and its vice versa without surfactant (NH2SO4 1%). Increase in grain yield due to treatments effect over weed free ranged from 23.8 to 37.0 per cent. B:C ratio also followed same path as in case of grain yield.

BACKGROUND:The study seeks to investigate the impact of Food and Drug Administration's black box warning (BBW) on the use of atypical antipsychotics (AAP) and nonantipsychotic psychotropic alternatives in noninstitutionalized elderly population diagnosed with dementia.METHOD:The Medical Expenditure Panel Survey (2004 through 2007) was utilized as the data source. Medication use in elderly patients (≥65 years) was defined as taking at least 1 medication for dementia. We performed a statistical comparison of prewarning (2004-2005) and postwarning (2006-2007) periods with respect to AAP and nonantipsychotic psychotropic use to examine the impact of labeling changes.RESULTS:A bivariate analysis did not yield statistically significant change in either the AAP or nonantipsychotic psychotropic use, pre- versus postwarning. However, multivariate logistic-regression analyses revealed greater odds for antidementia (odds ratio [OR] = 1.976, P = .0195) and benzodiazepine (OR = 3.046, P = .0227) medication use in postwarning period compared to the prewarning period.CONCLUSION:The regulatory warnings and labeling changes regarding off-label use of AAPs in dementia treatment showed minimal impact on their actual use in noninstitutionalized populations. A corresponding increase in the use of nonantipsychotic psychotropics explains that BBW might have resulted in a compensatory shift in favor of benzodiazepines and antidementia medications. Additional research should be conducted to examine the long-term impact of BBW on antipsychotic prescribing changes.