Antimicrobial resistance and phylogenetic relatedness of extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli in peridomestic rats (Rattus norvegicus and Rattus tanezumi) linked to city areas and animal farms in Hong Kong.

Extended-spectrum ß-lactamase-producing Escherichia (E.) coli (ESBL-EC) in the clinical setting have emerged as a major threat to public and animal health. Wildlife, including Rattus spp. may serve as reservoirs and spreaders of ESBL-EC in the environment. Peridomestic rats are well adapted to living in proximity to humans and animals in a variety of urban and agricultural environments and may serve as sentinels to identify variations of ESBL-EC within their different habitats. In this study, a set of 221 rats (Rattus norvegicus, R. tanezumi, R. andamanensis, and Niviventer huang) consisting of 104 rats from city areas, 44 from chicken farms, 52 from pig farms, and 21 from stables of horse-riding schools were screened for ESBL-EC. Overall, a total of 134 ESBL-EC were isolated from the caecal samples of 130 (59%) rats. The predominant blaESBL genes were blaCTX-M-14, blaCTX-M-15, blaCTX-M-55, and blaCTX-M-65. Phylogenetic analysis revealed a total of 62 sequence types (STs) and 17 SNP clusters. E. coli ST10 and ST155 were common to ESBL-EC from city areas and chicken farms, and ST44 were found among ESBL-EC from city areas and pig farms. Extra-intestinal pathogenic E. coli (ExPEC) ST69, ST131 and ST1193 were found exclusively among rats from city areas, and avian pathogenic E. coli (APEC) ST177 was restricted to ESBL-EC originating from chicken farms. Phylogenetic analysis showed that the populations of rodent ESBL-EC from city areas, chicken farms and pig farms were genetically different, suggesting a certain degree of partitioning between the human and animal locations. This study contributes to current understanding of ESBL-EC occurring in rats in ecologically diverse locations.

Prevalence and characterization of antimicrobial-resistant Escherichia coli in chicken meat from wet markets in Hong Kong.

Given the close contact between animals, animal products, and consumers in wet markets, fresh meat products are considered a potential source and disseminator of antimicrobial-resistant (AMR) bacteria near the end of the food chain. This cross-sectional study was conducted to estimate the prevalence of select AMR-E. coli in fresh chicken meat collected from wet markets in Hong Kong and to determine target genes associated with the observed resistance phenotypes. Following a stratified random sampling design, 180 fresh half-chickens were purchased from 29 wet markets across Hong Kong in 2022 and immediately processed. After incubation, selective isolation was performed for extended-spectrum ß-lactamase producing (ESBL), carbapenem-resistant (CRE), and colistin-resistant (CSR) E. coli. The bacterial isolates were identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Disc Diffusion was used to determine the susceptibility of ESBL- and CRE-E. coli isolates. The broth microdilution method was used to determine the minimum inhibitory concentration of CSR-E. coli. Targeted resistance genes were then detected by PCR. The prevalence of ESBL-E. coli and CSR-E. coli were estimated at 88.8% (95% CI: 83.4-93.1%) and 6.7% (95% CI: 3.5-11.4%), respectively. No CRE-E. coli isolate was detected. The blaCTX-M-1 gene was the most common ß-lactamase group in isolated E. coli (80%), followed by blaTEM (63.7%); no blaSHV gene was detected. Forty-five percent of the isolates had blaTEM and blaCTX-M-1 simultaneously. The mcr-1 gene was detected in all 12 CSR isolates. Of 180 meat samples, 59 were from Mainland China, and 121 were locally sourced. There was no statistically significant difference in the prevalence of ESBL- and CSR-E. coli between the two sources. Our findings can be used to inform food safety risk assessments and set the stage for adopting targeted control and mitigation measures tailored to the local wet markets.

Serotypes, Antimicrobial Resistance Profiles, and Virulence Factors of Salmonella Isolates in Chinese Edible Frogs (Hoplobatrachus rugulosus) Collected from Wet Markets in Hong Kong.

Salmonella is an important agent of gastrointestinal disease in humans. While livestock, such as cattle, poultry, and pigs, are well-recognised animal reservoirs of Salmonella, there is a lack of data on Salmonella in edible frogs, even though frog meat is a popular food worldwide. In this study, 103 live edible Chinese frogs (Hoplobatrachus rugulosus) were collected from wet markets throughout Hong Kong. After euthanasia, faeces or cloacal swabs were examined for Salmonella. Overall, Salmonella spp. were isolated from 67 (65%, CI: 0.554-0.736) of the samples. The serotypes included S. Saintpaul (33%), S. Newport (24%), S. Bareilly (7%), S. Braenderup (4%), S. Hvittingfoss (4%), S. Stanley (10%), and S. Wandsworth (16%). Many isolates were phylogenetically related. A high number of genes encoding for resistance to clinically relevant antimicrobials, and a high number of virulence determinants, were identified. Antimicrobial susceptibility testing (AST) identified multidrug resistance (MDR) in 21% of the isolates. Resistance to ampicillin, ciprofloxacin, nalidixic acid, and tetracycline was common. These results demonstrate that a high percentage of live frogs sold for human consumption in wet markets are carriers of multidrug-resistant Salmonella. Public health recommendations for handling edible frogs should be considered, to mitigate the risk of Salmonella transmission to humans.

Antimicrobial resistance in bacteria isolated from peridomestic Rattus species: A scoping literature review.

Rattus spp. may acquire and disseminate antimicrobial resistant bacteria or antimicrobial resistance (AMR) genes. We conducted a scoping review to synthesize available research findings on AMR in Rattus spp. and to describe the size and scope of available literature on AMR epidemiology in Rattus spp. The review was performed according to Preferred Reporting Items for Systematic Reviews and Meta-Analysis extension for Scoping Reviews (PRISMA-ScR). The search focused on scientific peer-reviewed publications focusing on AMR in peridomestic Rattus spp. The review was limited to publications in English available in PubMed, Web of Science and Scopus between 2000 and 2021. The results were summarized descriptively. Thirty-four studies conducted in twenty-one countries were included in this scoping review. Twelve bacterial species with AMR were identified with Escherichia coli and Staphylococcus aureus being the two most commonly reported. The resistant bacteria were isolated from species of peridomestic Rattus spp. in which R. norvegicus and R. rattus were the two most commonly studied. Rats were also found to carry multi-drug resistant (MDR) bacteria including extended-spectrum beta (ß)-lactamase (ESBL), methicillin-resistant Staphylococcus aureus (MRSA), colistin-resistant Enterobacteriaceae (CoRE), and vancomycin-resistant Enterococci (VRE). This scoping review suggests that peridomestic Rattus spp. can carry multiple antimicrobial resistant bacteria, indicating their potential to serve as reservoirs and spreaders of AMR thus posing a threat to human and animal health.

Complete Genome Sequence of the Extensively Drug-Resistant Extended-Spectrum ß-Lactamase-Producing Proteus mirabilis Isolate HK294, Obtained from Poultry Feces in Hong Kong.

Here, we report the complete genome sequence of Proteus mirabilis isolate HK294, recovered from pooled poultry feces in Hong Kong in 2022. The chromosome contained 32 antimicrobial resistance genes, including the extended-spectrum ß-lactamases blaCTX-M-65 and blaCTX-M-3. Almost all resistance genes were part of either an integrative conjugative element or a Tn7-like transposon.

Low Prevalence of SARS-CoV-2 Antibodies in Canine and Feline Serum Samples Collected during the COVID-19 Pandemic in Hong Kong and Korea.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has affected millions of people worldwide since its emergence in 2019. Knowing the potential capacity of the virus to adapt to other species, the serological surveillance of SARS-CoV-2 infection in susceptible animals is important. Hong Kong and Seoul are two of Asia's most densely populated urban cities, where companion animals often live in close contact with humans. Sera collected from 1040 cats and 855 dogs during the early phase of the pandemic in Hong Kong and Seoul were tested for SARS-CoV-2 antibodies using an ELISA that detects antibodies against the receptor binding domain of the viral spike protein. Positive sera were also tested for virus neutralizing antibodies using a surrogate virus neutralization (sVNT) and plaque reduction neutralization test (PRNT). Among feline sera, 4.51% and 2.54% of the samples from Korea and Hong Kong, respectively, tested ELISA positive. However, only 1.64% of the samples from Korea and 0.18% from Hong Kong tested positive by sVNT, while only 0.41% of samples from Korea tested positive by PRNT. Among canine samples, 4.94% and 6.46% from Korea and Hong Kong, respectively, tested positive by ELISA, while only 0.29% of sera from Korea were positive on sVNT and no canine sera tested positive by PRNT. These results confirm a low seroprevalence of SARS-CoV-2 exposure in companion animals in Korea and Hong Kong. The discordance between the RBD-ELISA and neutralization tests may indicate possible ELISA cross-reactivity with other coronaviruses, especially in canine sera.

The challenges of pig farming in Hong Kong: a study of farmers' perceptions and attitudes towards a pig health and production management service.

BACKGROUND: Pig farming in Hong Kong differs markedly from other places in the world, with a highly urbanised population, the majority of pigs being imported for slaughter, and limited on-farm veterinary support. Little is known about the barriers and attitudes of pig farmers in Hong Kong and their expectations of a new pig health and production management service provided by veterinarians. We collected qualitative and quantitative data to 1) describe pig farms, 2) identify barriers to pig farming in Hong Kong and 3) describe the perceptions of the new service. Thematic analysis was conducted to identify barriers and attitudes. RESULTS: Eight and nine out of 38 pig farmers agreed to participate in the qualitative and quantitative components, respectively. All farms were farrow-to-finish farms with a median of 2800 (range 950 to 7000) pigs per farm. Three themes were identified during the interview analysis and could be ranked based on their importance to the farmers: the regulatory environment (Theme 1), veterinary support structures (Theme 2), and the sustainability of the pig industry (Theme 3). Farmers expressed dissatisfaction with the regulation of the industry and veterinary services on offer within Hong Kong. However, farmers did note that the provision of a new pig health and production management service was as a positive development. The public perception of pig farming, market forces, and competition from mainland pig farmers have resulted in sustainability challenges for the industry. CONCLUSIONS: Farmers identified very specific local systems and challenges unique to pig farming in Hong Kong. The lack of veterinary support was one of these challenges and although a certain level of scepticism towards the new pig health and production service was expressed, farmers indicated their interest and listed areas where they would benefit from improved veterinary support. Prior experiences of veterinary services clouded farmers perceptions of the usefulness of a new service. To be successful in this environment, clear communication about the goals, role and limitations of the new on farm service is crucial, as is the alignment with the needs of farmers. Despite the small sample size, the qualitative methodology used allows us to assume that these themes give a general idea of what Hong Kong farmers' concerns and attitudes are.

Is there a higher risk of exposure to Coxiella burnetii for pre-clinical veterinary students?

Coxiella burnetii is globally distributed but evidence of zoonotic transmission in the Caribbean region is scarce. The bacterium presence is suspected on the Caribbean island of St. Kitts. The risk of exposure of veterinary students was reported in other regions of the world but is not documented in the Caribbean region. The present study aimed to evaluate the risk of exposure to C. burnetii for pre-clinical veterinary students (mostly coming from the U.S.) attending an island-based veterinary school. A cross-sectional study was conducted to compare incoming and outgoing veterinary students' seroprevalence. Serology was performed using indirect immunofluorescence assay to test Coxiella burnetii Phase I and Phase II immunoglobulins M and G. Background data were gathered using a standardized questionnaire. A parallel study enrolled veterinary school employees in the same university. Of the 98 participants (48 incoming and 50 outgoing students), 41 (41.8%, 95 %CI: 31.9-52.2) were seropositive to C. burnetii. There was no significant difference between the two groups (45.8% for incoming vs. 38.0% for outgoing students) (p = 0.4). No risk factors (demographic, animal handling practices or background) were significantly more reported in the seropositive group. In the employee study, the seroprevalence was high with 8/15 seropositives (53.3%, 95 %CI: 26.6-78.7). Pre-clinical veterinary students do not have a higher risk of exposure to C. burnetii by attending the veterinary school in St. Kitts, but they are highly exposed before arrival on the island (seroprevalence of 45.8%). Most of these participants had experience with animals either through farming or previous veterinary technician employment. This indicates a high exposure in the U.S. young population aiming to become veterinarians. There is an urgent need to increase C. burnetii surveillance in animals and humans to apply relevant prevention and control measures, including recommendations for vaccination of students and professionals at risk.

Neurological disease suspected to be caused by tick-borne encephalitis virus infection in 6 horses in Switzerland.

BACKGROUND: Reports on acute tick-borne encephalitis virus (TBEV) infections with signs of neurologic disease in horses are limited. OBJECTIVES: To describe the epidemiological, clinical, and laboratory findings of suspected acute TBEV infections in 6 horses. ANIMALS: Six horses originating from TBEV endemic regions of Switzerland were presented to equine hospitals with acute onset of neurologic disease between 2011 and 2019. METHODS: Retrospective case series. Horses with acute onset of signs of neurologic disease that were subjected to clinical and microbiological examinations to rule out infectious diseases affecting the central nervous system. RESULTS: All horses exhibited acute signs of neurologic disease including ataxia and proprioceptive deficits. Horses tested positive for TBEV using virus neutralization test and samples were further tested for TBEV-specific IgM. The presence of TBEV-specific IgM antibodies was confirmed in 5 horses (cases 1-5, Laboratory Unit [LU] values ranging from 30 to 56). One horse (case no. 6) with an LU value just below the test threshold (LU = 22.3) was also included under the hypothesis that the horse was transitioning from acute to chronic infection. All horses originated from areas where humans with confirmed tick-borne encephalitis reported to have been bitten by ticks. CONCLUSIONS AND CLINICAL IMPORTANCE: Acute TBEV infection should be a differential diagnosis in horses with signs of neurologic disease and originating from TBEV endemic areas. The establishment of harmonized diagnostic criteria would help to overcome the diagnostic challenges associated with TBEV and other Flavivirus infections in horses.

Serotype Diversity and Antimicrobial Resistance Profile of Salmonella enterica Isolates From Freshwater Turtles Sold for Human Consumption in Wet Markets in Hong Kong.

Chelonians are recognized as a source of human salmonellosis through direct contact or consumption of their meat. Freshwater turtles sold for food are widely available in wet markets in Asia. In this pilot study, 50 turtles belonging to three species were randomly sampled from wet markets throughout Hong Kong. The turtles were humanely euthanised and their feces or the colon were sampled for Salmonella culture. The Salmonella isolates obtained were serotyped and examined for phenotypic antimicrobial resistance and the presence of antimicrobial resistance genes. The study reports a high prevalence (42%, 95% CI: 29.4-55.8) and considerable serotype diversity of Salmonella among turtles sold in wet markets. The most common among the 11 serotypes isolated were S. Oranienburg and S. Thompson, which have been reported in turtles previously. The serotype S. Manhattan is reported in chelonians for the first time. Resistance to streptomycin and chloramphenicol was common, despite the latter being banned from aquaculture in mainland China since 2002. Resistance against fluoroquinolones and third-generation cephalosporins which represent first-line treatment options for salmonellosis was also observed. The multidrug-resistance gene cfr is identified for the first time in Salmonella. This is a worrying finding as it indicates an expansion of the cfr reservoir and potential horizontal spread to other bacteria. The results of this study emphasize the need for close surveillance of Salmonella from turtles sold as food and better regulation of turtle farming to safeguard public health and improve animal welfare.

The screening for anticoagulant rodenticide gene VKORC1 polymorphism in the rat Rattus norvegicus, Rattus tanezumi and Rattus losea in Hong Kong.

Anticoagulants are a major component of rodenticides used worldwide, which function by effectively blocking the vitamin K cycle in rodents. The rat Vitamin K epoxide Reductase Complex (VKORC) subunit 1 is the enzyme responsible for recycling vitamin K, and five substitution mutations (Tyr139Cys, Tyr139Ser, Tyr139Phe and Leu128Gln and Leu120Gln) located in the VKORC1 could result in resistance to anticoagulant rodenticides. This study carried out a VKORC1-based survey to estimate the anticoagulant rodenticide resistance in three Rattus species (R. losea, R. norvegicus, and R. tanezumi) collected in Hong Kong. A total of 202 rats captured in Hong Kong between 2017 and 2021 were analysed. Sequencing of molecular marker cytochrome c oxidase subunit 1 (COX1) was carried out to assist the species identification, and the identities of 52 lesser ricefield rats (R. losea), 81 common rats (R. norvegicus) and 69 house rats (R. tanezumi) were confirmed. Three VKORC1 exons were amplified from individuals by PCR followed by Sanger sequencing. A total of 47 R. tanezumi (68.1%) contained Tyr139Cys mutation in VKORC1 gene, and half of them were homozygous. None of the collected R. losea and R. norvegicus were detected with the five known substitutions leading to anticoagulant rodenticides resistance, and previously undescribed missense mutations were revealed in each species. Whole genome sequencing was further carried out on some individuals, and single nucleotide polymorphisms (SNPs) were also identified in the introns. This is the first study investigating the situation of anticoagulant rodenticide resistance in the rats collected in Hong Kong. Given that the efficacy of rodenticides is crucial for effective rodent management, regular genetic testing as well as population genomic analyses will be required to both monitor the situation and understand the adaption of different rat haplotypes for integrated pest management. Susceptibility tests for individual rodenticides should also be conducted regularly to assess their effectiveness on local species.

Quality Assessment of Day-Old Chickens on the Broiler Farms of Hong Kong.

The present study aimed to investigate the quality of newly hatched broiler chicks delivered to Hong Kong (imported or local), and to develop tailored recommendations to improve their management. During 2019-2021, 70 batches (34 imported from mainland China and 36 local) of one/three day old chicks on 11 broiler farms were studied. From each batch, 23 or 24 chicks (1647 in total) were assessed for abnormalities in appearance, navel, crop filling, dehydration, body weight, and length uniformity. Chicks were sacrificed, and yolk sac residues in three day old chicks were measured. Maternally derived antibody levels against Newcastle disease virus (NDV), infectious bursal disease virus (IBDV), and avian reovirus (ARV) were measured in all chicks using an enzyme-linked immunosorbent assay (ELISA). The proportion of abnormal navel in most batches is high (median: 59%), ranging between 0 and 100%. The average length of chicks within batches ranges between 16.3 and 20.7 cm, and their average weights are 31-38.5 g upon delivery to the farms. On average, imported batches have a higher body weight and length than their local counterparts. The average yolk-free weight varies between 45 and 55 g, which is significantly lower in local batches (33-43 g). The mean antibody titers against NDV and ARV are higher in imported batches than in the local ones. In contrast, the mean antibody titer against IBDV is significantly lower in the imported batches. Concerning the overall lower quality of local chicks compared to the imported batches, establishing a well-managed broiler breeder farm and a hatchery in Hong Kong is highly recommended to enhance the health and productivity of the local broiler chicken industry.

Evaluation of Vaccination Strategy Against Rabies in Hong Kong Macaques.

The objectives of this study were to assess the serological response to rabies vaccination in Hong Kong macaques and provide evidence-based recommendations for the vaccination interval implemented by the Government of Hong Kong. An inactivated rabies vaccine was administered subcutaneously to captured macaques under a mass sterilization program in Hong Kong. Blood samples from the animals were collected in a 2015 field survey and stored in -80°C freezer. In 2021, the frozen sera from vaccinated animals were prepared and tested for antibodies against the rabies virus using a commercial blocking enzyme-linked immunosorbent assay (ELISA) test. Sixty-five samples were available from the vaccinated macaques that had received at least one dose of the vaccine between 2008 and 2015. The interval between the first vaccination and blood sampling ranged from 21 to 2,779 days (median: 990). Only five macaques had a second vaccination record at the sampling time, all with high antibody levels. Among the remaining macaques, 77% (46/60) were positive for rabies antibodies. No specific association was observed between the post-vaccination period and the antibody titer of these macaques, and no adverse reactions were reported. Although the precise level of protection against a potential challenge with the rabies virus cannot be ascertained, the vaccination elicited rabies antibodies in 87% (21/24) of the macaques tested within 2.5 years of their first vaccination. Our findings indicate the potential benefits of the current vaccination strategy to protect the population from rabies and consequential mandatory culling of all macaques if a natural infection occurs.

Does having a cat in your house increase your risk of catching COVID-19?

Due to the zoonotic origin of SARS-Coronavirus 2 (SARS-CoV-2), the potential for its transmission from humans back to animals and the possibility that it might establish ongoing infection pathways in other animal species has been discussed. Cats are highly susceptible to SARS-CoV-2 and were shown experimentally to transmit the virus to other cats. Infection of cats has been widely reported. Domestic cats in COVID-19-positive households could therefore be a part of a human to animal to human transmission pathway. Here, we report the results of a qualitative risk assessment focusing on the potential of cat to human transmission in such settings. The assessment was based on evidence available by October 2021. After the introduction of SARS-CoV-2 to a household by a human, cats may become infected and infected cats may pose an additional infection risk for other members of the household. In order to assess this additional risk qualitatively, expert opinion was elicited within the framework of a modified Delphi procedure. The conclusion was that the additional risk of infection of an additional person in a household associated with keeping a domestic cat is very low to negligible, depending on the intensity of cat-to-human interactions. The separation of cats from humans suffering from SARS-CoV-2 infection should contribute to preventing further transmission.

A tool for rapid assessment of wildlife markets in the Asia-Pacific Region for risk of future zoonotic disease outbreaks.

Decades of warnings that the trade and consumption of wildlife could result in serious zoonotic pandemics have gone largely unheeded. Now the world is ravaged by COVID-19, with tremendous loss of life, economic and societal disruption, and dire predictions of more destructive and frequent pandemics. There are now calls to tightly regulate and even enact complete wildlife trade bans, while others call for more nuanced approaches since many rural communities rely on wildlife for sustenance. Given pressures from political and societal drivers and resource limitations to enforcing bans, increased regulation is a more likely outcome rather than broad bans. But imposition of tight regulations will require monitoring and assessing trade situations for zoonotic risks. We present a tool for relevant stakeholders, including government authorities in the public health and wildlife sectors, to assess wildlife trade situations for risks of potentially serious zoonoses in order to inform policies to tightly regulate and control the trade, much of which is illegal in most countries. The tool is based on available knowledge of different wildlife taxa traded in the Asia-Pacific Region and known to carry highly virulent and transmissible viruses combined with relative risks associated with different broad categories of market types and trade chains.

Expert opinion on livestock antimicrobial usage indications and patterns in Denmark, Portugal and Switzerland.

We aimed at describing antimicrobial usage patterns throughout livestock production cycles, and comparing them across three countries from Northern, Central and Southern Europe. Given the difficulties to collect such detailed usage data, an expert opinion was deemed the most appropriate study design. This study provides new insights into the time periods and indications for which specific antimicrobial substances are used in different livestock sectors. Veterinary experts (n=67) from different livestock sectors (broilers, pigs, dairy cattle and veal/fattening calves) and countries (Denmark, Portugal and Switzerland) replied to a questionnaire focusing on the time periods in the production cycle when antimicrobial substances were administered, and the respective indications for treatment. Our results showed that for several antimicrobials, between-country and within-country variations exist regarding the temporal distributions of treatments and indications for use. These differences were also true for several critically important antimicrobials, which is a matter of concern. Furthermore, differences between countries were also evident regarding the antimicrobial substances licensed. Based on our results, it is recommended to establish and promote treatment guidelines, invest in the prevention of diseases during critical moments of the production cycle and target undifferentiated use of antimicrobials. Moreover, discrepancies between countries should be further investigated to better understand the factors underlying the identified patterns and to distinguish prudent from non-prudent use. The results can inform decision-making with the aim to foster antimicrobial prudent use in the veterinary setting and, therefore, protect public health from the threat of antimicrobial resistance.