Prapas Patchanee

Genetic relatedness and molecular characterization of multidrug resistant Acinetobacter baumannii isolated in central Ohio, USA

BackgroundOver the last decade, nosocomial infections due to Acinetobacter baumannii have been described with an increasing trend towards multidrug resistance, mostly in intensive care units. The aim of the present study was to determine the clonal relatedness of clinical isolates and to elucidate the genetic basis of imipenem resistance.MethodsA. baumannii isolates (n = 83) originated from two hospital settings in central Ohio were used in this study. Pulsed-field gel electrophoresis genotyping and antimicrobial susceptibility testing for clinically relevant antimicrobials were performed. Resistance determinants were characterized by using different phenotypic (accumulation assay for efflux) and genotypic (PCR, DNA sequencing, plasmid analysis and electroporation) approaches.ResultsThe isolates were predominantly multidrug resistant (>79.5%) and comprised of thirteen unique pulsotypes, with genotype VII circulating in both hospitals. The presence of blaOXA-23 in 13% (11/83) and ISAba1linked blaOXA-66 in 79.5% (66/83) of clinical isolates was associated with high level imipenem resistance. In this set of OXA producing isolates, multidrug resistance was bestowed by blaADC-25, class 1 integron-borne aminoglycoside modifying enzymes, presence of sense mutations in gyrA/parC and involvement of active efflux (with evidence for the presence of adeB efflux gene).ConclusionThis study underscores the major role of carbapenem-hydrolyzing class D β-lactamases, and in particular the acquired OXA-23, in the dissemination of imipenem-resistant A. baumannii. The co-occurrence of additional resistance determinant could also be a significant threat.

Tracking Salmonella Contamination in Various Watersheds and Phenotypic and Genotypic Diversity

Salmonella enterica is an important foodborne pathogen, and contamination of surface and ground water that may result from various human activities, such as animal production and urbanization, may contribute to the public health burden. The aims of this study was to determine the sources of Salmonella contamination in four different types of watersheds and to assess the relative contribution of multidrug-resistant strains. Eighty-six water samples collected from four different watershed systems, including those impacted by swine production (n = 12), residential/industrial (n = 34), crop agriculture (n = 12), and forestry (n = 28), were cultured for Salmonella and further characterized by serotyping, antimicrobial susceptibility testing, and pulsed-field gel electrophoresis genotyping. Salmonella prevalence was high in all four watersheds: residential/industrial area (58.8%), forestry (57.1%), crop agriculture (50%), and swine production (41.7%). Majority of the Salmonella isolates (87.1%) were pansusceptible. Multidrug resistance up to eight antimicrobials (R-type: AmStTeAxChCeKmGm) was detected in water samples that originated from swine production systems only. Serovars identified included Anatum, Gaminara, and Inverness (18.3% each) and Muenchen and Newport (8.7% each), Bredeny (7.6%), and Montevideo (6.8%). Pulsed-field gel electrophoresis analysis indicated genotypic relatedness among Salmonella recovered from residential/industrial and forestry-associated watersheds (genotypic cluster types A, C, D, E, F, G, H, and J), sites with relatively close geographic proximity. Swine-production-associated isolates were distinctly different from the others (genotypic cluster types B and I), corroborating the phenotypic findings. Overall, the findings suggest that all the various watersheds, including natural forest, remain important contributors of Salmonella contamination. While swine-production-associated water samples were not found to have a disproportionately high prevalence, it was the most important reservoir of multidrug-resistant strains.

The Global One Health Paradigm: Challenges and Opportunities for Tackling Infectious Diseases at the Human, Animal, and Environment Interface in Low- Resource Settings

Zoonotic infectious diseases have been an important concern to humankind for more than 10,000 years. Today, approximately 75% of newly emerging infectious diseases (EIDs) are zoonoses that result from various anthropogenic, genetic, ecologic, socioeconomic, and climatic factors. These interrelated driving forces make it difficult to predict and to prevent zoonotic EIDs. Although significant improvements in environmental and medical surveillance, clinical diagnostic methods, and medical practices have been achieved in the recent years, zoonotic EIDs remain a major global concern, and such threats are expanding, especially in less developed regions. The current Ebola epidemic in West Africa is an extreme stark reminder of the role animal reservoirs play in public health and reinforces the urgent need for globally operationalizing a One Health approach. The complex nature of zoonotic diseases and the limited resources in developing countries are a reminder that the need for implementation of Global One Health in low-resource settings is crucial. The Veterinary Public Health and Biotechnology (VPH-Biotec) Global Consortium launched the International Congress on Pathogens at the Human-Animal Interface (ICOPHAI) in order to address important challenges and needs for capacity building. The inaugural ICOPHAI (Addis Ababa, Ethiopia, 2011) and the second congress (Porto de Galinhas, Brazil, 2013) were unique opportunities to share and discuss issues related to zoonotic infectious diseases worldwide. In addition to strong scientific reports in eight thematic areas that necessitate One Health implementation, the congress identified four key capacity-building needs: (1) development of adequate science-based risk management policies, (2) skilled-personnel capacity building, (3) accredited veterinary and public health diagnostic laboratories with a shared database, and (4) improved use of existing natural resources and implementation. The aim of this review is to highlight advances in key zoonotic disease areas and the One Health capacity needs.

Characterization of Multidrug-Resistant Salmonella enterica Serovar Heidelberg Isolated from Humans and Animals

Salmonella enterica serovar Heidelberg has been recognized as one of the most common serovar associated with foodborne infections in the United States. It is also frequently isolated from nonhuman sources and has increasingly shown resistance to various antimicrobial agents. The present study was undertaken to identify the predominant antimicrobial resistance phenotypes and genotypes of Salmonella Heidelberg (n = 95) isolates of human, swine, and turkey origin. Antimicrobial susceptibility was done using Kirby-Bauer disk diffusion method with a panel of 12 antimicrobials. Pulsed-field gel electrophoresis genotyping was used to determine the diversity of the isolates. The antimicrobial resistance genes and carriage of Class 1 and 2 integrons were determined by polymerase chain reaction. All Salmonella Heidelberg isolates from swine were resistant to one or more of the antimicrobials tested and the majority (73.3%) showed multidrug resistance to streptomycin, tetracycline, and kanamycin (R-type: StTeKm). About 80% of the Salmonella Heidelberg isolates of human origin were pan-susceptible, however, one isolate showed multidrug resistance to 10 of 12 antimicrobials tested. Among the multidrug-resistant (MDR) Salmonella Heidelberg isolates, Class 1 integrons with variable sizes of 1.2 to 1.5 kb were detected in six isolates (three each) from humans and swine. DNA sequencing revealed that Class 1 integrons of both human and swine origin carried a gene encoding aminoglycoside adenyltransferase (aadA). Resistance genes identified in other loci include aphA1-Iab, strA, bla(TEM), and tetA (B). Both human and swine MDR strains of Salmonella Heidelberg carried the resistance phenotypes on self-transferable plasmids. Dendrogram analysis of pulsotypes indicated possible clonality of Salmonella Heidelberg between isolates of human and swine origin. The findings in this study indicate the increasing significance of swine as reservoirs of emerging MDR serovars, such as MDR Salmonella Heidelberg, is of public health significance.

A cross-sectional study of hepatitis E virus infection in healthy people directly exposed and unexposed to pigs in a rural community in northern Thailand.

A cross‐sectional study of the association between occupational pig exposure and hepatitis E virus (HEV) infection in adult pig farmers and the general population who were not directly exposed to pigs was conducted in Nan Province, Thailand, from November 2010 to April 2011. All participants were interviewed to provide information on their job history, eating habits and other potential confounders. The prevalence of anti‐HEV immunoglobulin G antibodies (IgG) among 513 subjects was 23.0%. Hand washing with water and soap was associated with a lower seroprevalence of HEV infection, whereas living in an area with frequent flooding (OR 1.64, 95% CI: 1.00–2.68) and consuming internal pig organs more than twice per week (OR 3.23, 95%CI: 1.15–9.01) were both associated with a higher seroprevalence of anti‐HEV IgG. There was no association between HEV seroprevalence and frequent, direct occupational pig contact.

Prevalence, quantitative load, and antimicrobial resistance of Campylobacter spp. from broiler ceca and broiler skin samples in Thailand

This study was conducted to determine the prevalence of Campylobacter spp. in broiler flocks by testing cecal contents at slaughter and to detect and quantify Campylobacter on broiler carcass skin samples of the corresponding slaughter batches, to determine antimicrobial resistance patterns of the Campylobacter isolates, and to genotype selected Campylobacter jejuni isolates using multilocus sequence typing analysis. Ninety-eight broiler flocks were included in the study. Intact ceca were randomly taken at the time of evisceration throughout a slaughter batch to detect Campylobacter spp. at the broiler flock level and one whole carcass per slaughter batch was taken for the detection of Campylobacter spp. on broiler skin. The prevalences of Campylobacter spp. in broiler ceca and broiler skin samples were 11.2% (11/98) and 51% (50/98), respectively. Even though most Campylobacter-positive broiler skin samples were contaminated with only up to 230 most probable number per gram, a substantial share (13.3%) showed very high Campylobacter numbers on the broiler skin samples (most probable number = ∞; lower confidence limit T(0) 580/g). From 32 C. jejuni and Campylobacter coli isolates tested, the highest antimicrobial resistance rates were found for ciprofloxacin (81.2%), followed by tetracycline (40.6%), ampicillin (31.2%), and erythromycin (9.4%). All tested strains were sensitive to gentamicin. By multilocus sequence typing analysis, a total of 9 different sequence types were identified among 16 C. jejuni isolates. Campylobacter jejuni isolated from cecal content and carcass skin of the same farm or slaughter batch showed corresponding allelic profiles. Our data suggest that intense cross-contamination during the slaughter process led to a strong increase of Campylobacter prevalence on broiler skin compared with the prevalence in broiler ceca. To reduce Campylobacter prevalences on broiler skin, on-farm biosecurity measures need to be accompanied by control measures at the slaughterhouse to reduce fecal contamination of broiler skin and to minimize cross-contamination.

Occurrence and characterization of livestock-associated methicillin-resistant Staphylococcus aureus in pig industries of northern Thailand.

This study was conducted to determine the prevalence of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) in pigs, farm workers, and the environment in northern Thailand, and to assess LA-MRSA isolate phenotypic characteristics. One hundred and four pig farms were randomly selected from the 21,152 in Chiang Mai and Lamphun provinces in 2012. Nasal and skin swab samples were collected from pigs and farm workers. Environmental swabs (pig stable floor, faucet, and feeder) were also collected. MRSA was identified by conventional bacterial culture technique, with results confirmed by multiplex PCR and multi locus sequence typing (MLST). Herd prevalence of MRSA was 9.61% (10 of 104 farms). Among pigs, workers, and farm environments, prevalence was 0.68% (two of 292 samples), 2.53% (seven of 276 samples), and 1.28% (four of 312 samples), respectively. Thirteen MRSA isolates (seven from workers, four from environmental samples, and two from pigs) were identified as Staphylococcal chromosomal cassette mec IV sequences type 9. Antimicrobial sensitivity tests found 100% of the MRSA isolates resistant to clindamycin, oxytetracycline, and tetracycline, while 100% were susceptible to cloxacillin and vancomycin. All possessed a multidrug-resistant phenotype. This is the first evidence of an LA-MRSA interrelationship among pigs, workers, and the farm environment in Thailand.

A cross-sectional study of Hepatitis e virus infection in pigs in different-sized farms in Northern Thailand

Pigs are an important reservoir of hepatitis E virus (HEV) in many countries throughout the world. We evaluated the association between farm size and presence of serum antibodies against HEV, as well as other risk factors for infection in pigs raised in Nan Province, Thailand in a cross-sectional study. The sampling frame was a total-population census of all pig herds, stratified into three classes of the farm size according to criteria developed by the Nan provincial livestock health office. One-eighth of all pigs in each farm were sampled randomly. All pig-farm owners were interviewed to elicit information on general characteristics of their farms, biosecurity and hygienic procedures, and farm management. We obtained sera and fecal samples from 879 pigs to test for antibodies to HEV and HEV RNA. Odds ratios (OR) and 95% confidence intervals (CI) for risk factors for HEV seroprevalence were estimated by multivariate logistic regression. The overall prevalence of anti-HEV immunoglobulin G antibodies was 9.9%. Pigs studied from medium-sized farms had a higher HEV seroprevalence than those from larger farms (adjusted OR 4.95, 95% CI: 1.79, 13.70). Factors associated with HEV seropositivity included feeding pigs with agro-industrial byproducts, having veterinarians on farms, and presence of other pig farms within 100 m. Twenty-five (2.9%) of 875 sampled pig stools were positive for HEV RNA. Phylogenetic analysis revealed that all HEV isolates clustered to HEV genotype 3.

Comparative Phenotypic and Genotypic Characterization of Salmonella spp. in Pig Farms and Slaughterhouses in Two Provinces in Northern Thailand

Salmonella spp. are an important group of bacterial zoonotic pathogens which can cause acute food-borne diseases in humans. Pork products are the main source of salmonellosis, but the origins and transmission routes of the disease have not been clearly determined. The purpose of this study was to characterize Salmonella spp. isolated in pig production lines both from pig farms and from slaughterhouses in Chiang Mai and Lamphun provinces in northern Thailand. The study focuses on the association among serotypes, antimicrobial resistance patterns and Pulse Field Gel Electrophoresis (PFGE) patterns to investigate possible sources of infection and to provide information which could help strengthen salmonellosis control programs in the region. A total of 86 strains of Salmonella comprising five majority serotypes were identified. Antibiotic resistance to tetracycline was found to be the most prevalent (82.56%) followed by ampicillin (81.40%) and streptomycin (63.95%). Seven clusters and 28 fingerprint-patterns generated by PFGE were identified among strains recovered from various locations and at different times, providing information on associations among the strains as well as evidence of the existence of persistent strains in some areas. Study results suggest that Salmonella control programs should be implemented at slaughterhouse production lines, including surveillance to insure good hygiene practices, in addition to regular monitoring of large populations of farm animals.

Comparative phenotypic and genotypic analyses of Salmonella Rissen that originated from food animals in Thailand and United States.

Salmonella enterica serovar Rissen has been recognized as one of the most common serovar among humans and pork production systems in different parts of the world, especially Asia. In the United States, this serovar caused outbreaks but its epidemiologic significance remains unknown. The objectives of this study were to compare the phenotypic (antimicrobial susceptibility) and genotypic attributes of Salmonella Rissen isolated in Thailand (Thai) and the United States (US). All the Thai isolates (n = 30) were recovered from swine faecal samples. The US isolates (n = 35) were recovered from swine faecal samples (n = 29), cattle (n = 2), chicken (n = 2), dog (n = 1) and a ready‐to‐eat product (n = 1). The antimicrobial susceptibility of isolates was determined using the Kirby‐Bauer disk diffusion method with a panel of 12 antimicrobials. Pulse‐field gel electrophoresis (PFGE) was used to determine the genotypic diversity of isolates. All Thai isolates showed multidrug resistance (MDR) with the most frequent antibiotic resistance shown against ampicillin (100%), sulfisoxazole (96.7%), tetracycline (93.3%), streptomycin (90%) and chloramphenicol (30%). About half of the isolates of USA origin were pan‐susceptible and roughly 30% were resistant to only tetracycline (R‐type: Te). Salmonella Rissen isolated from Thailand and the USA in this study were found to be clonally unrelated. Genotypic analyses indicated that isolates were clustered primarily based on the geographic origin implying the limited clonality among the strains. Clonal relatedness among different host species within the same geography (USA) was found. We found genotypic similarity in Thai and US isolates in few instances but with no epidemiological link. Further studies to assess propensity for increased inter‐regional transmission and dissemination is warranted.