Animal Fecal Samples Research Articles

Quantitative PCR versus metagenomics for monitoring antibiotic resistance genes: balancing high sensitivity and broad coverage.

The widespread occurrence of clinically relevant antibiotic resistance within humans, animals, and environment motivates the development of sensitive and accurate detection and quantification methods. Metagenomics and quantitative PCR (qPCR) are amongst the most used approaches. In this study, we aimed to evaluate and compare the performance of these methods to screen antibiotic resistance genes in animal faecal, wastewater, and water samples. Water and wastewater samples were from hospital effluent, different treatment stages of two treatment plants, and of the receiving river at the discharge point. The animal samples were from pig and chicken faeces. Antibiotic resistance gene coverage, sensitivity, and usefulness of the quantitative information were analyzed and discussed. While both methods were able to distinguish the resistome profiles and detect gradient stepwise mixtures of pig and chicken faeces, qPCR presented higher sensitivity for the detection of a few antibiotic resistance genes in water/wastewater. In addition, the comparison of predicted and observed antibiotic resistance gene quantifications unveiled the higher accuracy of qPCR. Metagenomics analyses, while less sensitive, provided a markedly higher coverage of antibiotic resistance genes compared to qPCR. The complementarity of both methods and the importance of selecting the best method according to the study purpose are discussed.

Extending TF-Test to the diagnosis of gastrintestinal parasites of humans and animals.

Parasitic species inhabit the intestinal tract of humans and animals, and they can cause damages whose severity depends on several factors related to the parasite and the host. Immunocompromised individuals are more likely to develop severe forms of parasitic infestation. The gastrointestinal parasites diagnosis is performed mainly by the parasitological examination of feces, which consists of the visualization and identification of the parasitic structures eliminated along with the feces. These exams have low sensitivity and a large number of impurities, which can impair the result of the diagnosis. In order to improve the diagnosis quality, a new parasitological technique called Three Fecal Test (TF‑Test) was developed. Thus, we have approached in this study new techniques to diagnose gastrointestinal parasites from human and animal fecal samples. These techniques originate from adaptations and improvements of the TF‑Test technique protocol. From these operational changes it has been possible to carry out the diagnosis of several human and animal parasites, with satisfactory results.

Portable Differential Detection of CTX-M ESBL Gene Variants, blaCTX-M-1 and blaCTX-M-15, from Escherichia coli Isolates and Animal Fecal Samples Using Loop-Primer Endonuclease Cleavage Loop-Mediated Isothermal Amplification.

Cefotaximase-Munich (CTX-M) extended-spectrum beta-lactamase (ESBL) enzymes produced by Enterobacteriaceae confer resistance to clinically relevant third-generation cephalosporins. CTX-M group 1 variants, CTX-M-1 and CTX-M-15, are the leading ESBL-producing Enterobacteriaceae associated with animal and human infection, respectively, and are an increasing antimicrobial resistance (AMR) global health concern. The blaCTX-M-1 and blaCTX-M-15 genes encoding these variants have an approximate nucleotide sequence similarity of 98.7%, making effective differential diagnostic monitoring difficult. Loop-primer endonuclease cleavage loop-mediated isothermal amplification (LEC-LAMP) enables rapid real-time multiplex pathogen detection with single-base specificity and portable on-site testing. We have developed an internally controlled multiplex CTX-M-1/15 LEC-LAMP assay for the differential detection of blaCTX-M-1 and blaCTX-M-15. Assay analytical specificity was established using a panel of human, animal, and environmental Escherichia coli isolates positive for blaCTX-M-1 (n = 18), blaCTX-M-15 (n = 35), and other closely related blaCTX-Ms (n = 38) from Ireland, Germany, and Portugal, with analytical sensitivity determined using probit regression analysis. Animal fecal sample testing using the CTX-M-1/15 LEC-LAMP assay in combination with a rapid DNA extraction protocol was carried out on porcine fecal samples previously confirmed to be PCR-positive for E. coli blaCTX-M. Portable instrumentation was used to further analyze each fecal sample and demonstrate the on-site testing capabilities of the LEC-LAMP assay with the rapid DNA extraction protocol. The CTX-M-1/15 LEC-LAMP assay demonstrated complete analytical specificity for the differential detection of both variants with sensitive low-level detection of 8.5 and 9.8 copies per reaction for blaCTX-M-1 and blaCTX-M-15, respectively, and E. coli blaCTX-M-1 was identified in all blaCTX-M positive porcine fecal samples tested. IMPORTANCE CTX-M ESBL-producing E. coli is an increasing AMR public health issue with the transmission between animals and humans via zoonotic pathogens now a major area of interest. Accurate and timely identification of ESBL-expressing E. coli CTX-M variants is essential for disease monitoring, targeted antibiotic treatment and infection control. This study details the first report of portable diagnostics technology for the rapid differential detection of CTX-M AMR markers blaCTX-M-1 and blaCTX-M-15, facilitating improved identification and surveillance of these closely related variants. Further application of this portable internally controlled multiplex CTX-M-1/15 LEC-LAMP assay will provide new information on the transmission and prevalence of these CTX-M ESBL alleles. Furthermore, this transferable diagnostic technology can be applied to other new and emerging relevant AMR markers of interest providing more efficient and specific portable pathogen detection for improved epidemiological surveillance.

Gut Microbiota Alterations in Trace Amine-Associated Receptor 9 (TAAR9) Knockout Rats

Trace amine-associated receptors (TAAR1-TAAR9) are a family of G-protein-coupled monoaminergic receptors which might have great pharmacological potential. It has now been well established that TAAR1 plays an important role in the central nervous system. Interestingly, deletion of TAAR9 in rats leads to alterations in the periphery. Previously, we found that knockout of TAAR9 in rats (TAAR9-KO rats) decreased low-density lipoprotein cholesterol levels in the blood. TAAR9 was also identified in intestinal tissues, and it is known that it responds to polyamines. To elucidate the role of TAAR9 in the intestinal epithelium, we analyzed TAAR9-co-expressed gene clusters in public data for cecum samples. As identified by gene ontology enrichment analysis, in the intestine, TAAR9 is co-expressed with genes involved in intestinal mucosa homeostasis and function, including cell organization, differentiation, and death. Additionally, TAAR9 was co-expressed with genes implicated in dopamine signaling, which may suggest a role for this receptor in the regulation of peripheral dopaminergic transmission. To further investigate how TAAR9 might be involved in colonic mucosal homeostasis, we analyzed the fecal microbiome composition in TAAR9-KO rats and their wild-type littermates. We identified a significant difference in the number of observed taxa between the microbiome of TAAR9-KO and wild-type rats. In TAAR9-KO rats, the gut microbial community became more variable compared with the wild-type rats. Furthermore, it was found that the family Saccharimonadaceae, which is one of the top 10 most abundant families in TAAR9-KO rat feces, is almost completely absent in wild-type animal fecal samples. Taken together, these data indicate a role of TAAR9 in intestinal function.

The Antibacterial Effects of Cocktail and Single Forms of Lytic Phages Belonging to Podoviridae and Myoviridae Families from Sewage against Shigella sonnei and Shigella flexneri.

Diarrhea caused by bacterial pathogens such as Shigella spp. is one of the prominent public health concerns. The evolution of vast antibiotic resistance by these pathogens, leading to failure in the infections eradication, has made an impetus to seek and develop novel approaches. Recently, some alternative therapies such as phage therapy have been investigated. Bacteriophages are viruses that target specific bacterial species. The objective of this study was to assess the therapeutic effect of phages obtained from hospital sewage against Shigella sonnei (S. sonnei) ATCC® 9290 and S. flexneri ATCC 12022 standard and clinical strains. Four various lytic bacteriophages were isolated from animal fecal and sewage samples and propagated using S. sonnei and S. flexneri as host organisms. The phages' morphology was determined using transmission electron microscopy (TEM). The lytic potential and host specificity of isolated phages were evaluated using double layer plaque assay and spot test. Moreover, bacterial turbidity values were evaluated in coculture with phages in the Luria Bertani (LB) medium for 24 hours at time intervals of 30 min. Phage cocktails (Shs1, Shs2, Shf1, and Shf2) exhibited higher antimicrobial activity than single phage application against S. sonnei and S. flexneri standard strains. The phages belonged to Podoviridae and Myoviridae families according to TEM-assisted morphological features analysis. In addition, the phages exhibited host specificity using the spot test against 18 Shigella clinical isolates. In this study, phage cocktail of Podoviridae and Myoviridae families from sewage conferred substantial antibacterial effects against S. sonnei and S. flexneri. However, single phage effects were unstable in the LB coculture. Moreover, the phages had host specificity using the spot test performed against Shigella spp. clinical isolates.

Low doses and short duration of prednisolone administration in guinea pigs experimentally infected with Schistosoma haematobium: Histopathology of liver and lungs

In our previous study, administration of 5 mg prednisolone for five days pre-Schistosoma haematobium infection in guinea pigs increased susceptibility and produced pathological reactions in the liver and bladder. Since corticosteroids can suppress granuloma formation, maturation, and size, this study sought to investigate if prednisolone given at low doses and short duration can produce granulomatous lesions in the tissues of guinea pigs experimentally infected with S. haematobium. Guinea pigs were shared into six groups: group I and II were the immunosuppressed-infected guinea pigs (I0.5 and I1.5- 20 animals each), group III was the unimmunosuppressed-infected guinea pigs (UI- 20 animals), and group IV, V and VI were the immunosuppressed-uninfected and normal guinea pigs (D0.5, D1.5, and normal- 10 animals each). Prednisolone was given in doses of 0.5 mg/kg and 1.5 mg/kg to the different groups, a day before infection and on day 5 post-infection. The infected groups were subcutaneously injected with 250–300 S. haematobium cercariae. Screening for S. haematobium eggs in urine and fecal samples of animals, and quantitative analysis for leukocyte and red blood cell (RBC) counts in urine samples of guinea pigs began nine weeks post-infection (WPI). Guinea pigs were killed, perfused, worms recovered and sections of the liver, lungs, and bladder excised for histopathological examination at 6, 8, 11, 14 and 16 WPI. S. haematobium eggs were only seen in urine samples of I1.5 at 15 and 16WPI. Although the parasite eggs were seen in fecal samples of all infected guinea pigs from 9WPI, those of UI were sparse and took longer time to hatch. High leukocyte counts were seen in all immunosuppressed groups at 6WPI, which returned to normal levels in D1.5 and D0.5 at 16WPI. At 16WPI, significant numbers of leukocyte and RBC counts were seen in urine samples of I1.5. The immunosuppressed-infected groups had significant numbers of mature and total worm loads than UI group (p > 0.05). However, the worm burden of I1.5 was higher than I0.5 at 14WPI and 16WPI. Non-granulomatous lesions were only recorded in the liver sections of the immunosuppressed-infected animals and in lung sections of UI and I1.5 guinea pigs. Liver lesions seen were hepatocyte degeneration; necrosis; Kupffer cell involvements as hyperplasia, phagocytosis, proliferation; hyperaemia and haemorrhage, and mononuclear leukocyte infiltration. Lung lesions seen in I1.5 at 11-16WPI were hemosiderin depositions and hyperaemia, emphysema and atelectasis, and mononuclear leukocyte infiltrations while in UI, emphysema and mononuclear leukocyte infiltration were seen only at 16WPI. In the immunosuppressed-infected groups, composite liver lesion scores showed that peak lesion severity was at 8WPI and 11WPI in I1.5 and I0.5, respectively. However, there was no significant difference (p = 0.105) in composite liver lesion scores of I1.5 and I0.5. Lung lesion score of UI at 16WPI was significantly higher (p > 0.05) than that of I1.5. Findings from this study show that even at low doses and short duration of administration, corticosteroids can only increase susceptibility of guinea pigs but cannot improve its suitability as experimental models of S. haematobium infection.

Helminth infections of captive animals and management practices at the University of Ilorin Zoo, North-Central, Nigeria

This study investigated the helminth parasites of animals and management practices at the University of Ilorin Zoological garden. Faecal samples were collected and analysed according to standard parasitological techniques while information on management practices was obtained using a structured questionnaire. The overall prevalence of helminths was 20.7%. Three parasites namely, Ascaris spp., Trichostrongylus spp. and Enterobius spp. were identified. Of the 92 animal faecal samples examined, common jackal (21.1%), crested porcupine (42.1%), Lioness (15.8%), and mongoose (21.1%) were positive (p<0.05). Only mammals were positive for intestinal helminth while Aves were not infected (p>0.05). Animals fed three times a week had higher helminth infection (57.9%) when compared with animals fed daily (42.1%). Cages cleaned daily had more helminth parasite contamination (78.9%) compared to those cleaned weekly (21.1%, p<0.05). Also, animals dewormed on monthly basis harboured fewer intestinal helminths (5.3%), compared to those dewormed every three months (78.9%, p>0.05). Surprisingly, animals in enclosures with cemented floors had the highest helminth infection (78.9%) compared to those living in enclosures with uncemented floor (p<0.05). The overall prevalence of helminths is low in this study; therefore, zoo workers should take more cognizance of the routes of parasite transmission, since the faecal-oral route is the primary means through which animals acquired infection. There is a need for improved management practices to curb the spread of helminth infection. This will invariably ensure the sustainability of zoological gardens.

Zoo animal manure as an overlooked reservoir of antibiotic resistance genes and multidrug-resistant bacteria.

Animal fecal samples collected in the summer and winter from 11 herbivorous animals, including sable antelope (SA), long-tailed goral (LTG), and common eland (CE), at a public zoo were examined for the presence of antibiotic resistance genes (ARGs). Seven antibiotics, including meropenem and azithromycin, were used to isolate culturable multidrug-resistant (MDR) strains. The manures from three animals (SA, LTG, and CE) contained 104-fold higher culturable MDR bacteria, including Chryseobacterium, Sphingobacterium, and Stenotrophomonas species, while fewer MDR bacteria were isolated from manure from water buffalo, rhinoceros, and elephant against all tested antibiotics. Three MDR bacteria-rich samples along with composite samples were further analyzed using nanopore-based technology. ARGs including lnu(C), tet(Q), and mef(A) were common and often associated with transposons in all tested samples, suggesting that transposons carrying ARGs may play an important role for the dissemination of ARGs in our tested animals. Although several copies of ARGs such as aph(3')-IIc, blaL1, blaIND-3, and tet(42) were found in the sequenced genomes of the nine MDR bacteria, the numbers and types of ARGs appeared to be less than expected in zoo animal manure, suggesting that MDR bacteria in the gut of the tested animals had intrinsic resistant phenotypes in the absence of ARGs.

Plasmid and chromosomal copies of blaCMY-2 mediate resistance to third-generation cephalosporins in Escherichia coli from food animals in China

The use of antimicrobials in food animals is the major determinant for the propagation of resistant bacteria in the animal reservoir. The objective of this study was to investigate the presence and distribution of third-generation cephalosporin (3GC) -resistant and plasmid-mediated AmpC (pAmpC)-producing Escherichia coli isolated from food animals in Southern China. In total, 744 3GC-resistant and 40 blaCMY-2-positive E. coli strains were recovered from 1656 food animal fecal samples across five rearing regions. The blaCMY-2 genes were located on IncC, IncFIB or IncI1 type plasmids in 12 E. coli isolates. In the other 22 isolates, S1-PFGE and hybridization analyses revealed that the blaCMY-2 gene was chromosomally located and demonstrated a high prevalence of the chromosomally encoded blaCMY-2 gene in E. coli. Plasmid stability and growth curve experiments demonstrated that IncI1, IncC and IncFIB plasmids can exist stably in the host bacteria and with a low growth burden and may be the reason these plasmids can be widely disseminated in breeding environments. Whole genome sequencing indicated that ISEcp1 and IS1294 played important roles in blaCMY-2 transfer to both plasmids and the chromosome. Our study confirmed that blaCMY-2 mediated resistance of food animal-derived E. coli to 3GC and highlights the urgent need for appropriate monitoring programmes.

Distribution, genetic diversity, and zoonotic significance of Blastocystis subtypes in pet dogs

The common intestinal protist, Blastocystis sp., is a highly frequent parasitic agent in human and animal fecal samples, globally. The current study evaluated the prevalence of this parasite and involved subtypes (STs) in examined pet dogs in Shiraz, southwestern Iran. A number of 100 fresh fecal specimens were randomly collected from pet dogs referred to the veterinary clinics and pet shops in Shiraz during July 2021-January 2022. The genomic DNA was extracted from fecal samples and a conventional PCR was performed. Next, sequencing of the positive samples was done based on the small subunit ribosomal rRNA gene. The results showed that 5 % (5 out of 100) of the examined samples were positive for Blastocystis. As anticipated, three known Blastocystis subtypes with zoonotic concern, ST1 (60 %, n = 3), ST2 (20 %, n = 1) and ST3 (20 %, n = 1), were characterized. Based on phylogenetic analysis, these subtypes showed 100% identity with their reference counterparts. In conclusion, the zoonotic subtypes found in pet dogs in Shiraz can pose a drastic challenge to their pet owners and other domestic animals. Albeit the role of dogs as a potential natural reservoir of Blastocystis sp. stays unknown, it appears that these animals could represent possible reservoirs of zoonotic transmission of Blastocystis sp. Still, there is paucity of data on the epidemiology of Blastocystis and its subtypes in dogs in Iran.

First Molecular Study and Sequencing of Cryptosporidium spp. Isolated from Human, Cattle and Sheep in Wasit Province, Iraq

The current study aimed to investigate Cryptosporidium spp. that infect humans and some animal species as well as to compare among various isolates obtained. A total of 96 human stool samples and 96 animal fecal samples (48 for cattle and 48 for sheep) were collected and examined by using Nested PCR technique followed by DNA sequencing analysis through phylogenetic tree to know the parasite species. The whole infection rate in human was 36.4% (35/96), while in cattle was 22.9% (11/48) and in sheep was 14.5% (7/48) by nested polymerase chain reaction technique. The findings of DNA sequencing based on 18s rRNA gene (654 bp) showed the presence of three Cryptosporidium spp. which namely C. hominis, C. parvum, and C. bovis, all of them have identity 99 %, Cryptosporidium parvum was the most widespread species being detected in 60% (9/15), followed by C. bovis 33.33% (5/15), and C. hominis 6.66% (1/15). This is the first report revealed the molecular sequencing among human, cattle and sheep in Wasit province, Iraq and the role of livestock (cattle, sheep) in zoonotic transmission of the cryptosporidiosis.

Genome and virulome characterization of Escherichia coli encoding multiple antimicrobial resistance genes isolated from commercial dairy operations

The aim of this study was to describe the genome sequences of 12 Escherichia coliisolates that encode antimicrobial resistance genes (ARGs) and were isolated from composite dairy animal faecal samples. The isolates were recovered between 2013 and 2014 from a larger study evaluating the presence of resistance on dairy operations in Pennsylvania, USA. The draft genomes were sequenced on an Illumina NextSeq 500 platform and were assembled using SPAdes. In total, 69 ARGs were identified, and these were categorized into 20 unique genes conferring resistance to eight classes of antimicrobials. In order of decreasing frequency of detection, these classes were aminoglycosides, tetracyclines, sulphonamides, β-lactams, phenicols, trimethoprim, fosfomycin, and macrolide-lincosamide-streptogramin B (MLS). Among the resistant isolates, 11 sequence types (ST) were identified, with ST86 detected twice. Although pathogenicity was not evaluated in vivo, multiple genomes encoded virulence factors involved in severe gastrointestinal and extraintestinal infections, including a single Shiga-toxigenic isolate. The results of this study demonstrate the presence of antimicrobial-resistant E. coli in dairy animal faeces encoding virulence factors involved in severe human infections.

Evaluation of crAssphage as a human-specific microbial source-tracking marker in the Republic of Korea.

CrAssphage is a novel and by far the most abundant bacteriophage in the human gut and has been proposed as a human-specific microbial source tracking (MST) marker. However, its global use as a human-specific MST marker requires validation in more extensive regions. The purpose of this study was to evaluate the specificity and abundance of the human-specific MST marker crAssphage with PCR and RT-PCR assays in human and animal feces in Korea. The prevalence of crAssphage was confirmed in 94 human feces samples (subjects: 19 to 45years old) and 56 animal feces samples (from birds, raccoons, squirrels, weasels, deer, wild boars, hares, cats, and dogs). CrAssphage showed sensitivity of 0.39 and specificity of 1.00 in Korea, with a sequencing analysis showing that genotype II was dominant at 71.9%. The quantitative analysis showed that crAssphage is sufficiently abundant in human feces given the high concentration range of 4.26 to 8.25 log gene copies (GC)/ng in human feces. In conclusion, this study confirmed the crAssphage as a specific and abundant MST marker with which to identify human fecal contamination in Korea.

Molecular Identification of Bacillus Isolated from Korean Water Deer (Hydropotes inermis argyropus) and Striped Field Mouse (Apodemus agrarius) Feces by Using an SNP-Based 16S Ribosomal Marker.

Simple SummaryWildlife is a great concern because of its free-ranging movements. They carry bacterial zoonoses in their feces, such as Bacillus species. In this study, we developed a 16S Bacillus species-specific 16S ribosomal RNA (rRNA) molecular marker for species identification. For discrimination of genetically similar members of Bacillus cereus group, including Bacillus cereus, B. anthrax, and B. thuringiensis, a single nucleotide polymorphism (SNP)-based marker was developed. We altered an artificial base at the 3′-end of SNP sites in each SNP-based 16S rRNA primer sequence to improve the ability of SNP-based primers to bind the PCR template sequence, thereby improving the allele-specific detection of target B. cereus strains. SNP analysis in high-quality DNA sequences could facilitate identification and discrimination of closely related bacterial species.Ambiguous, heterogeneous, endospore-forming Bacillus species, notably Bacillus cereus, often produce fatal toxins that threaten human health. We identified Bacillus from wild animal fecal samples (n = 80), including the Korean water deer (n = 25) and striped field mouse (n = 55). Using traditional culture-based methods, 25 animal fecal samples (31.25%; 25/80) were found to be positive for Bacillus species, whereas using molecular techniques, 19 samples (23.75%; 19/80) were found to be positive for the same. In addition, we designed a Bacillus species-specific 16S ribosomal RNA (rRNA) gene marker and utilized it to identify 19 samples by means of PCR amplification and sequencing, using at least one colony from the 19 Bacillus positive samples. The recovered sequences were matched to sequences of three Bacillus species (B. cereus, B. amyloliquefaciens, and B. megaterium) from the GenBank database. Moreover, the phylogenetic tree generated in this study established specific clades for the Bacillus group. In addition, to differentiate between B. cereus, B. anthracis, and B. thuringiensis, we designed a single nucleotide polymorphism (SNP)-based primer by identifying SNPs in the alignment of 16S rRNA gene sequences of B. cereus group strains. The SNPs were used to design primer sets for discrimination between highly similar species from the B. cereus group. The study could be used in surveillance of agricultural fresh-produce-associated Bacillus outbreaks, for accurate identification of each Bacillus species, and in the development of control measures.

Development of a qPCR assay for the detection of naturalized wastewater E. coli strains.

We recently demonstrated the presence of naturalized populations of Escherichia coli in municipal sewage. We wanted to develop a quantitative polymerase chain reaction (qPCR) assay targeting the uspC-IS30-flhDC marker of naturalized wastewater E. coli and assess the prevalence of these naturalized strains in wastewater. The limit of detection for the qPCR assay was 3.0 × 10-8 ng of plasmid DNA template with 100% specificity. This strain was detected throughout the wastewater treatment process, including treated effluents. We evaluated the potential of this marker for detecting municipal sewage/wastewater contamination in water by comparing it to other human and animal markers of fecal pollution. Strong correlations were observed between the uspC-IS30-flhDC marker and the human fecal markers Bacteroides HF183 and HumM2, but not animal fecal markers, in surface and stormwater samples. The uspC-IS30-flhDC marker appears to be a potential E. coli-based marker for human wastewater contamination.

Risk factors for third-generation cephalosporin-resistant and extended-spectrum β-lactamase-producing Escherichia coli carriage in domestic animals of semirural parishes east of Quito, Ecuador.

Extended-spectrum β-lactamase (ESBL)-producing and other antimicrobial resistant (AR) Escherichia coli threaten human and animal health worldwide. This study examined risk factors for domestic animal colonization with ceftriaxone-resistant (CR) and ESBL-producing E. coli in semirural parishes east of Quito, Ecuador, where small-scale food animal production is common. Survey data regarding household characteristics, animal care, and antimicrobial use were collected from 304 households over three sampling cycles, and 1195 environmental animal fecal samples were assessed for E. coli presence and antimicrobial susceptibility. Multivariable regression analyses were used to assess potential risk factors for CR and ESBL-producing E. coli carriage. Overall, CR and ESBL-producing E. coli were detected in 56% and 10% of all fecal samples, respectively. The odds of CR E. coli carriage were greater among dogs at households that lived within a 5 km radius of more than 5 commercial food animal facilities (OR 1.72, 95% CI 1.15-2.58) and lower among dogs living at households that used antimicrobials for their animal(s) based on veterinary/pharmacy recommendation (OR 0.18, 95% CI 0.04-0.96). Increased odds of canine ESBL-producing E. coli carriage were associated with recent antimicrobial use in any household animal (OR 2.69, 95% CI 1.02-7.10) and purchase of antimicrobials from pet food stores (OR 6.83, 95% CI 1.32-35.35). Food animals at households that owned more than 3 species (OR 0.64, 95% CI 0.42-0.97), that used antimicrobials for growth promotion (OR 0.41, 95% CI 0.19-0.89), and that obtained antimicrobials from pet food stores (OR 0.47, 95% CI 0.25-0.89) had decreased odds of CR E. coli carriage, while food animals at households with more than 5 people (OR 2.22, 95% CI 1.23-3.99) and located within 1 km of a commercial food animal facility (OR 2.57, 95% CI 1.08-6.12) had increased odds of ESBL-producing E. coli carriage. Together, these results highlight the complexity of antimicrobial resistance among domestic animals in this setting.

Clonal and plasmid-mediated dissemination of environmental carbapenem-resistant Enterobacteriaceae in large animal breeding areas in northern China

The emergence and dissemination of carbapenem-resistant Enterobacteriaceae (CRE) constitute a major global health problem. The environment plays an important role in the dissemination of CRE, but large-scale studies on CRE in groundwater environments in animal breeding areas are scarce. The aim of this study was to investigate CRE occurrence and environmental transmission of carbapenem resistance genes in large animal breeding areas in northern China. In total, 280 well water and 102 animal feces samples in large animal breeding areas in six counties from the two provinces Inner Mongolia and Shandong in China, were screened for CRE. A total of 39 CRE were isolated and characterized with next-generation sequencing. 5.3% of well water samples were contaminated with CRE. The well water in chicken farms had the highest number of detections of CRE (15.9%). More than half of the isolates carried closely related, conjugative IncX3 plasmids with blaNDM-genes from multiple geographic areas, indicating that this kind of plasmid plays an important role in dissemination of carbapenem resistance determinants. The clonal expansion of various CRE isolates in well water and animal feces were demonstrated; clonally related CRE were isolated from different wells within the same county, from different counties in the same province, and even from different provinces. In addition to harboring various ARGs, two closely related K. pneumoniae belonging to ST11 isolated from well water carried genetic hypervirulence determinants on a virulence plasmid, highlighting the potential health risk posed by further dissemination of this strain. These findings suggest that groundwater may be an underappreciated reservoir and source of dissemination of CRE, from which resistance genes may disseminate among different bacterial strains and over large geographic distances. Further research and multi-sectorial monitoring, with a “One health” perspective, is urgently needed to investigate the need for interventions aimed at preventing CRE dissemination.

Drinking quality assessment of well water with respect to Escherichia coli contamination from different sources#

The present study was undertaken to assess the drinking water quality of well water with respect to Escherichia coli contamination from different sources. A total of 138 drinking water samples contaminated with coliforms above 10 MPN Index per 100 mL formed the study material. A cross sectional survey was conducted for the sanitary evaluation of drinking water along with inspection of household wells and surroundings. A retrospective investigation was carried out to assess the contamination of E. coli-positive well water samples. The associated sources for E. coli contamination like, 93 human stool samples, 84 animal faecal samples, 25 bird cloacal swabs, 138 soil and 27 drainage water samples were also collected from the premises of the respective households in order to identify the source contaminating the well water. All the samples were subjected to isolation and identification of E. coli by conventional culture technique followed by molecular confirmation using polymerase chain reaction (PCR). Statistical analysis using Pearson’s chi square test revealed highly significant association between the occurrence of E. coli in drinking water and sources viz., human stool, animal faeces and soil samples (p ≤ 0.01). The unassociated sources of E. coli contamination in drinking water exhibited a highly significant association with the increasing distance of well from the septic tank as well as with the presence of linings in wells and animals in the households. Hence, the study conducted for the identification of sources of E. coli contamination of drinking water signifies the importance of revised construction rules to protect the wells from contaminating sources and the need of regular and effective water treatment protocols.

Agritourism and Kidding Season: A Large Outbreak of Human Shiga Toxin-Producing Escherichia coli O157 (STEC O157) Infections Linked to a Goat Dairy Farm-Connecticut, 2016.

The objective of this study was to determine sources of Shiga toxin-producing Escherichia coli O157 (STEC O157) infection among visitors to Farm X and develop public health recommendations. A case-control study was conducted. Case-patients were defined as the first ill child (aged <18 years) in the household with laboratory-confirmed STEC O157, or physician-diagnosed hemolytic uremic syndrome with laboratory confirmation by serology, who visited Farm X in the 10 days prior to illness. Controls were selected from Farm X visitors aged <18 years, without symptoms during the same time period as case-patients. Environment and animal fecal samples collected from Farm X were cultured; isolates from Farm X were compared with patient isolates using whole genome sequencing (WGS). Case-patients were more likely than controls to have sat on hay bales at the doe barn (adjusted odds ratio: 4.55; 95% confidence interval: 1.41–16.13). No handwashing stations were available; limited hand sanitizer was provided. Overall, 37% (29 of 78) of animal and environmental samples collected were positive for STEC; of these, 62% (18 of 29) yielded STEC O157 highly related by WGS to patient isolates. STEC O157 environmental contamination and fecal shedding by goats at Farm X was extensive. Farms should provide handwashing stations with soap, running water, and disposable towels. Access to animal areas, including animal pens and enclosures, should be limited for young children who are at risk for severe outcomes from STEC O157 infection. National recommendations should be adopted to reduce disease transmission.

Gastrointestinal parasites in wild and exotic animals from a Zoobotanical Park in Northeast of Brazil

One of the obstacles for ex situ conservation of wild and exotic animals are the diseases that affect them in captivity and, among them, the endoparasitoses, which are very frequent. The objective of this study is to report the occurrence of endoparasites in the fecal samples of animals from the Arruda Câmara Zoobotanical Park, at João Pessoa, Paraiba State, Brazil, as well as to identify significant statistical differences between the percentages of each parasite species found. To this end, 66 fecal samples were obtained from 50 species of animals including mammals, birds, and reptiles. Statistical differences between the percentages of each parasite species were obtained using the binomial test at 5% significance level. A total of 54.5% (36/66) of the fecal samples were positive (p=0.539), among which 80.5% (29/36) presented nematodes, followed by cestodes 19.4% (07/36), protozoans 13.9% (05/36), and mites 16.7% (06/36). This study reports for the first time the association of Balantidium sp. with Tapirus terrestris, Bertiella sp. with Alouatta caraya, Hymenolepis spp., and Aspiculuris spp. as spurious parasites in reptiles; and Entamoeba coli and Eimeria spp. with Iguana iguana in the northeastern part of the country. This study provides knowledge about some of the endoparasites that may occur in zoos in the northeastern region of the country, as well as in expanding the ecological data on wild and exotic animals.