Lifestyle behavior of physiotherapy students from Ecuador upon admission to higher education: a cross-sectional study.

Introduction: Admission to university has been identified as a period involving the adoption of unhealthy lifestyle behaviors. However, few studies have addressed the extent of this experience among Latin American university students. The aim of this study was twofold: first, to describe anthropometric variables, body composition, physical activity, sedentary behavior, sleep duration and quality, diet, and alcohol consumption in first-year students entering physiotherapy school at the Universidad de las Americas in Quito, Ecuador; second, to test differences in these variables between sexes. Methods: A total of 116 students were recruited. Sociodemographic variables, anthropometric indices, body composition, physical activity, sedentary behavior, sleep quality and duration, adherence to 24-hour movement guidelines, physical fitness (i.e., handgrip strength), diet, alcohol consumption, and smoking habits were evaluated. Results: A total of 50 male (43.1%) and 66 female (56.9%) students were assessed. Overall, the adherence to the 24-hour movement guidelines of the students upon admission to university was 8.6%. Conversely, 86 students (74.1%) did not meet any of the recommendations. Upon admission to university, only 8.6% of the students (female 2.6%; male 6%) met the overall 24-hour movement guidelines. Additionally, 82 students (70.7%) needed changes in diet quality, 81 students (69.8%) had significant sleep disturbances, and 22 students (18.9%) had harmful alcohol consumption. A greater proportion of males met all three 24-h movement recommendations (p = 0.025) than females did. In addition, females reported a greater percentage of occasional smokers (p = 0.025) and a greater prevalence of obesity (p < 0.001), a lower level of physical activity (p < 0.001), and a greater percentage of sleep disturbance (p < 0.001). Conversely, males reported greater waist circumference (p = 0.005), weight (p < 0.001), handgrip strength (p < 0.001), and a greater percentage of harmful alcohol consumption (p < 0.001). Discussion: Our study showed that upon admission to university, overall adherence to the 24-hour movement guidelines is low among university students. Additionally, a high percentage of students reported unhealthy lifestyle behaviors, with differences according to sex. Our findings could lead to the use of specific sex-specific strategies for the prevention and promotion of movement and lifestyle behaviors during this critical period.

Comparative genome analysis of colistin-resistant Escherichia coli harboring mcr isolated from rural community residents in Ecuador and Vietnam.

The spread of colistin-resistant bacteria among rural community residents of low- and middle-income countries is a major threat to community health. Although the mechanism of the spread of colistin-resistant bacteria in communities is unknown, geographic and regional characteristics may influence it. To elucidate the spread mechanism of colistin-resistant bacteria, we analyzed the genomes of colistin-resistant Escherichia coli isolated from Vietnam and Ecuador residents, which are geographically and socially different. Stool specimens of 139 and 98 healthy residents from Ecuador and Vietnam rural communities, respectively, were analyzed for colistin-resistant E. coli with mcr. Its prevalence in the residents of all the communities assessed was high and approximately equal in both countries: 71.8% in Ecuador and 69.4% in Vietnam. A phylogenetic tree analysis revealed that the sequence type of colistin-resistant E. coli was diverse and the major sequence types were different between the two countries. The location of mcr in the isolates showed that the proportion of chromosomal mcr was 35.1% and 8.5% in the Vietnam and Ecuador isolates, respectively. Most of these chromosomal mcr genes (75%-76%) had an intact mcr-transposon Tn6330. Contrastingly, the replicon types of the mcr-carrying-plasmids were diverse in both countries, but almost all belonged to IncI2 in Ecuador and IncX1/X4 in Vietnam. Approximately 26%-45% of these mcr-plasmids had other resistance genes, which also varied between countries. These results suggest that although the overall profile of the colistin-resistant E. coli isolates is diverse in these countries, the phylogenesis of the isolates and mcr-carrying plasmids has regional characteristics. Although the contributing factors are not clear, it is obvious that the overall profile of colistin-resistant bacteria dissemination varies between countries. Such different epidemic patterns are important for establishing country-specific countermeasures against colistin-resistant bacteria.

New Primers for Detection and Differentiation between Leishmania viannia and L. leishmania Subgenera by Polymerase Chain Reaction.

Background: Leishmania is the parasitic protozoan responsible for leishmaniases, a disease that can cause a range of cutaneous, mucosal, and visceral infections. Two subgenera L. Viannia and L. Leishmania are known to infect humans in the tropics and subtropics of the Americas. The aim of the present study was to develop a new pair of primers for the two subgenera and test in clinical samples. Methods: We designed two new pairs of primers for a PCR method from two conserved genes, cysteine proteinase B (cpb) and N-acetylglucosamine-6-phosfate deacetylase-like protein (nagA), as specific markers for those two respective subgenera. Primers were tested with 16 microscopical positive clinical samples from the Amazon region of Ecuador obtained in 2010-2020 period. Results: The cpb presented a band of 172 bp and the nagA a band of 300 bp, thus clearly differentiating L. viannia from L. leishmania. Additionally, primers identified and differentiated the clinical samples in the two subgenera. Conclusion: The new primers targeting different two genes and standardized in a PCR assay could identified and differentiated Leishmania parasites at subgenus level. This protocol could be used for Leishmania genus identification and diagnosis at the subgenus level and for determining the parasite's geographical distribution where different Leishmania subgenera are found in the same area.

A Fatal Case of Disseminated Histoplasmosis by Histoplasma capsulatum var. capsulatum Misdiagnosed as Visceral Leishmaniasis-Molecular Diagnosis and Identification.

Histoplasmosis is an endemic mycosis in the Americas. However, its diagnosis is challenging due to the complexity and limited availability of conventional laboratory techniques-antigen tests, culture, and staining. Microscopic preparations often confuse with other pathogens, such as Leishmania spp. The genus Histoplasma capsulatum comprises three varieties: var. capsulatum, var. duboissi, and var. farciminosum, which cannot be distinguished using conventional techniques. An infant from a tropical region of Ecuador was hospitalized for fever, bloody diarrhea, and anemia persisting for two months. Upon admission, he received antibiotics and immunosuppressants. Histopathological examination of the lymph nodes, intestines, and bone marrow aspirate reported the presence of Leishmania-like amastigotes, and treatment was initiated with meglumine antimoniate and conventional amphotericin B. However, subsequent analysis of samples using PCR and DNA sequencing identified H. capsulatum var. capsulatum but not Leishmania. Despite fluconazole and amphotericin B, the infant succumbed to the disease. The delay in clinical and laboratory diagnosis of histoplasmosis and the use of nonspecific and ineffective drugs such as fluconazole led to disease dissemination and, ultimately, death. Implementing molecular diagnosis and antigen tests in laboratories located in endemic regions and reference hospitals is crucial.

Case report: First report on human infection by tick-borne Babesia bigemina in the Amazon region of Ecuador.

Babesiosis is a protozoan disease acquired by the bite of different species of ticks. More than 100 Babesia spp. infect wild and domestic animals worldwide, but only a few have been documented to infect humans. Generally, babesiosis is asymptomatic in immunocompetent persons; however, in immunocompromised can be life-threatening. A 13-year-old boy from the Amazon region presented with a 3-month evolution of fever, chills, general malaise, and arthralgia accompanied by anemia and jaundice. In the last 4 years was diagnosed with chronic kidney failure. By nested-PCR using 18S RNA ribosomal gene as target and DNA sequencing, the phylogenetic analysis showed Babesia bigemina as the causative agent in the blood. Treatment with oral quinine plus clindamycin for six continuous weeks was effective with no relapse occurring during 12 months of follow-up. This is the second human case in Ecuador but the first caused by the zoonotic B. bigemina which confirms the existence of active transmission that should alert public health decision-making authorities on the emergence of this zoonosis and the need for research to determine strategies to reduce tick exposure.

The Development of a Multiplex PCR Assay for Fast and Cost-Effective Identification of the Five Most Significant Pathogenic Prototheca Species.

A multiplex PCR system (m-PCR) has been developed to accurately differentiate the five most important pathogenic Prototheca species, including the three species associated with infection in dairy cattle (P. ciferrii, P. blaschkeae, and P. bovis) and the two species associated with human infections (P. wickerhamii and P. cutis). The method is low-cost since it employs a simple "heat-shock" method in a TE buffer for DNA extraction. Furthermore, it requires only primers, a Taq polymerase, an agarose gel, and a molecular weight marker for identification. The method was based on published Prototheca cytochrome B sequences and was evaluated using reference strains from each of the five Prototheca species. The validity of the method was confirmed by identifying 50 strains isolated from milk samples. The specificity was tested in silico and with experimental PCR trials, showing no cross-reactions with other Prototheca species, as well as with bacteria, fungi, cows, algae, animals, or humans. The method could detect mixed infections involving two or three Prototheca species, providing a rapid test that delivers results within three hours.

Co-Harboring of Beta-Lactamases and mcr-1 Genes in Escherichia coli and Klebsiella pneumoniae from Healthy Carriers and Backyard Animals in Rural Communities in Ecuador.

Few studies have addressed drug resistance of Enterobacterales in rural communities in developing countries. This study aimed to determine the coexistence of extended-spectrum ß-lactamase (ESBL) and carbapenemase genes in Escherichia coli and Klebsiella pneumoniae strains carrying the mcr-1 gene in rural communities in Ecuador from healthy humans and their backyard animals. Sixty-two strains, thirty E. coli and thirty-two K. pneumoniae strains carrying the mcr-1 gene were selected from a previous study. PCR were performed for the presence of ESBLs and carbapenemase genes. The strains were further characterized, and the genetic relationship was studied with multi-locus sequencing typing (MLST) of seven housekeeping genes. Fifty-nine of the sixty-two mcr-1 isolates (95%) harbored at least on ß-lactam resistance gene. The most prevalent ESBL genes were the blaTEM genes (present in in 80% of the E. coli strains) and the blaSHV gene (present in 84% of the K. pneumoniae strains). MSLT analysis revealed 28 different sequence types (ST); 15 for E. coli and 12 for K. pneumoniae, with most ST never described in humans and animals. The coexistence of mcr-1 and ß-lactams resistant genes in E. coli and K. pneumoniae strains is alarming and threatens the efficacy of last-resort antibiotics. Our findings highlight backyard animals as a reservoir of mcr-1/ß-lactams resistant genes.

Colistin resistance in Escherichia coli and Klebsiella pneumoniae in humans and backyard animals in Ecuador.

Objective: Colistin is an antibiotic of last resort for treating serious Gram-negative bacterial infections. However, the misuse of colistin, especially as an animal growth promoter, has contributed to increasing antimicrobial resistance, mediated mainly through plasmid transfer of the mcr-1 gene. This study assessed the prevalence of phenotypic and molecular colistin resistance in Escherichia coli and Klebsiella pneumoniae in Ecuador in healthy humans and their chickens and pigs. Methods: Fecal samples were collected from humans and their chickens and pigs in two rural coastal and Amazon regions between April and August 2020. Gram-negative bacteria were isolated and identified using conventional techniques. Phenotypic resistance was determined using the broth microdilution technique, and the mcr-1 gene was detected using conventional polymerase chain reaction. Results: A total of 438 fecal samples were obtained from 137 humans, 147 pigs and 154 chickens. The prevalence of E. coli isolates was 86.3% (378/438) and K. pneumoniae, 37.4% (164/438). Overall, the mcr-1 gene was found in 90% (340/378) of E. coli isolates, with higher prevalences found in isolates from coastal regions (96.5%, 191/198), humans (95.6%, 111/116) and chickens (91.8%, 123/134); for K. pneumoniae, the gene was found in 19.5% (32/164) of isolates, with equal distribution between regions and hosts. Only four isolates, two E. coli and two K. pneumoniae, showed phenotypic resistance: mcr-1 was present in both E. coli strains but absent in the K. pneumoniae strains. Conclusions: Despite a low prevalence of phenotypic resistance to colistin, the high prevalence of the mcr-1 gene in E. coli is of concern. Ecuador's ban on using colistin in animal husbandry must be enforced, and continual monitoring of the situation should be implemented.

Whole-genome sequencing for surveillance of antimicrobial resistance in Ecuador: present and future implications.

Whole-genome sequencing is becoming the gold standard for pathogen characterization and offers considerable advantages for understanding the evolution and dissemination of new determinants of antimicrobial resistance. Despite the benefits of whole-genome sequencing for pathogen characterization, implementation costs and lack of expertise may limit its use by public health laboratories. This article reviews the advantages of whole-genome sequencing for pathogen characterization and the current status of the use of whole-genome sequencing for antimicrobial resistance surveillance in Ecuador. A roadmap is suggested for including whole-genome sequencing for pathogen characterization based on the needs of the health reference institutions through alliances with Ecuadorian universities. Establishing a partnership between public health institutions and academia would be valuable for clinicians, policy-makers, and epidemiologists who could then take reasonable measures in those areas and establish a basis for adapting One Health strategies to tackle antimicrobial resistance in Ecuador.

La secuenciación del genoma completo, que está pasando a ser el estándar de referencia para la caracterización de agentes patógenos, ofrece ventajas considerables para comprender la evolución y la diseminación de los nuevos determinantes de la resistencia a los antimicrobianos. Sin embargo, a pesar de los beneficios que genera, los costos de ejecución y la falta de experiencia pueden limitar su uso por parte de los laboratorios de salud pública. En este artículo se evalúan las ventajas de la secuenciación del genoma completo para la caracterización de agentes patógenos y el estado actual del uso de la secuenciación del genoma completo en la vigilancia de la resistencia a los antimicrobianos en Ecuador. Se propone una hoja de ruta para incluir la secuenciación del genoma completo para la caracterización de agentes patógenos según las necesidades de las instituciones de salud de referencia, lo que se haría por medio de alianzas con universidades ecuatorianas. Establecer una asociación entre las instituciones de salud pública y los círculos académicos sería sumamente valioso para los médicos, los responsables de las políticas y los epidemiólogos, que podrían adoptar medidas razonables en sus ámbitos y sentar una base para adaptar las estrategias de "Una salud" a fin de abordar la resistencia a los antimicrobianos en Ecuador.

O sequenciamento do genoma completo está se tornando o padrão ouro para a caracterização de patógenos e oferece vantagens consideráveis para a compreensão da evolução e disseminação de novos determinantes de resistência aos antimicrobianos. Apesar dos benefícios do sequenciamento do genoma completo para a caracterização de patógenos, os custos de implementação e a falta de especialização podem limitar seu uso pelos laboratórios de saúde pública. Este artigo analisa as vantagens do sequenciamento do genoma completo para a caracterização de patógenos e a situação atual do uso desta técnica para a vigilância da resistência aos antimicrobianos no Equador. Sugere-se um roteiro para incluir o sequenciamento de genomas completos para caracterização de patógenos com base nas necessidades das instituições de saúde de referência, por meio de alianças com universidades equatorianas. A criação de uma parceria entre instituições de saúde pública e entidades acadêmicas seria valiosa para clínicos, formuladores de políticas e epidemiologistas, que poderiam, assim, tomar medidas razoáveis nessas áreas e estabelecer uma base para adaptar estratégias de Saúde Única para combater a resistência aos antimicrobianos no Equador.

Diversity and prevalence of gastrointestinal helminths of free-roaming dogs on coastal beaches in Ecuador: Potential for zoonotic transmission.

Free-roaming dogs are highly exposed to a range of zoonotic parasites, including helminths, which can be transmitted to humans, particularly in rural tropical settings of developing countries. To evidence the diversity and prevalence of gastrointestinal helminths in the stools of free-roaming dogs on the public Pacific coastal beaches of Ecuador, a cross-sectional study was conducted from August 2021 to August 2022. The sampling beaches are located along the tropical Pacific coastal region. Stools were collected from the ground in containers with 10% formalin and processed using the Ritchie method; eggs were identified under a microscope. A total of 573 stools were examined from 20 beaches; the overall prevalence was 157 (27.4%) for one or more helminths. Ten parasites were identified, nine of which are potentially zoonotic. Ancylostoma spp. was the most prevalent (19.4%), followed by Toxocara spp. (7.2%). Trichuris spp., Dipylidium caninum, Diphyllobothrium spp., Capillaria spp., Dicrocoelium spp., Heterobilharzia americana, Hymenolepis spp. and Spirocerca spp. were also observed. Five of them are reported for the first-time infecting dogs in Ecuador. Hence, we evidenced that Ecuadorian beaches are highly contaminated with the dogs' zoonotic gastrointestinal helminths, posing a great risk to public health. Differences in the presence and prevalence were found in samples of tropical humid and dry climate zones. Therefore, based on our findings, we encourage the implementation of broad antiparasitic treatment and prevention strategies to reduce the zoonotic risk.

Colistin resistance in Escherichia coli and Klebsiella pneumoniae in humans and backyard animals in Ecuador

[ABSTRACT]. Objective. Colistin is an antibiotic of last resort for treating serious Gram-negative bacterial infections. However, the misuse of colistin, especially as an animal growth promoter, has contributed to increasing antimicrobial resistance, mediated mainly through plasmid transfer of the mcr-1 gene. This study assessed the prevalence of phenotypic and molecular colistin resistance in Escherichia coli and Klebsiella pneumoniae in Ecuador in healthy humans and their chickens and pigs. Methods. Fecal samples were collected from humans and their chickens and pigs in two rural coastal and Amazon regions between April and August 2020. Gram-negative bacteria were isolated and identified using conventional techniques. Phenotypic resistance was determined using the broth microdilution technique, and the mcr-1 gene was detected using conventional polymerase chain reaction. Results. A total of 438 fecal samples were obtained from 137 humans, 147 pigs and 154 chickens. The preva- lence of E. coli isolates was 86.3% (378/438) and K. pneumoniae, 37.4% (164/438). Overall, the mcr-1 gene was found in 90% (340/378) of E. coli isolates, with higher prevalences found in isolates from coastal regions (96.5%, 191/198), humans (95.6%, 111/116) and chickens (91.8%, 123/134); for K. pneumoniae, the gene was found in 19.5% (32/164) of isolates, with equal distribution between regions and hosts. Only four isolates, two E. coli and two K. pneumoniae, showed phenotypic resistance: mcr-1 was present in both E. coli strains but absent in the K. pneumoniae strains. Conclusions. Despite a low prevalence of phenotypic resistance to colistin, the high prevalence of the mcr-1 gene in E. coli is of concern. Ecuador’s ban on using colistin in animal husbandry must be enforced, and con- tinual monitoring of the situation should be implemented.

[RESUMEN]. Objetivo. La colistina es un antibiótico de último recurso para tratar infecciones graves por bacterias gramneg- ativas. Sin embargo, su uso indebido, especialmente para estimular el crecimiento animal, ha contribuido con el aumento de la resistencia a los antimicrobianos, mediada principalmente por la transferencia de plásmidos del gen mcr-1. En este estudio se evaluó la prevalencia de la resistencia fenotípica y molecular a la colistina de las bacterias Escherichia coli y Klebsiella pneumoniae en humanos sanos, sus pollos y cerdos en Ecuador. Métodos. Se recolectaron muestras fecales de humanos, así como de sus pollos y cerdos, en dos zonas rurales de la región costera y la región amazónica entre abril y agosto del 2020. Se aislaron las bacterias gramnegativas y se identificaron empleando técnicas convencionales. Se determinó la resistencia fenotípica mediante la técnica de microdilución en caldo y se detectó el gen mcr-1 con la técnica convencional de reac- ción en cadena de la polimerasa. Resultados. Se obtuvo un total de 438 muestras fecales de 137 humanos, 147 cerdos y 154 pollos. La preva- lencia de E. coli en las cepas aisladas fue del 86,3% (378/438) y la de K. pneumoniae, del 37,4% (164/438). En general, se detectó el gen mcr-1 en el 90% (340/378) de las cepas aisladas de E. coli y la mayor prevalencia encontrada fue en cepas aisladas de la región costera (96,5%, 191/198), humanos (95,6%, 111/116) y pollos (91,8%, 123/134); en el caso de K. pneumoniae, el gen se encontró en el 19,5% (32/164) de las cepas, con una distribución equitativa entre regiones y hospedadores. Únicamente cuatro cepas aisladas, dos de E. coli y dos de K. pneumoniae, mostraron resistencia fenotípica: el gen mcr-1 estaba presente en ambas cepas de E. coli y ausente en las cepas de K. pneumoniae. Conclusiones. Si bien hubo una baja prevalencia de resistencia fenotípica a la colistina, la alta prevalencia del gen mcr-1 en E. coli es preocupante. Es necesario hacer cumplir la prohibición del uso de colistina en la cría de animales en Ecuador, así como realizar un seguimiento continuo de la situación.

[RESUMO]. Objetivo. A colistina é um antibiótico de último recurso para o tratamento de infecções graves por bac- térias Gram-negativas. Entretanto, o uso indevido da colistina, principalmente como promotor de crescimento animal, tem contribuído para o aumento da resistência a antimicrobianos, principalmente por transferência horizontal do gene mcr-1 mediada por plasmídeos. Este estudo avaliou a prevalência de resistência fenotípica e molecular à colistina em Escherichia coli e Klebsiella pneumoniae no Equador em humanos hígidos e em galinhas e porcos por eles criados. Métodos. Entre abril e agosto de 2020, foram coletadas amostras de fezes de habitantes de duas regiões litorâneas e amazônicas do Equador e de galinhas e porcos por eles criados. Bactérias Gram-negativas foram isoladas e identificadas por meio de técnicas convencionais. A resistência fenotípica foi determinada pela técnica de microdiluição em caldo, e o gene mcr-1 foi detectado por reação em cadeia da polimerase convencional. Resultados. Foram obtidas 438 amostras fecais de 137 humanos, 147 suínos e 154 galinhas. A prevalência de isolados de E. coli foi de 86,3% (378/438), e de K. pneumoniae, 37,4% (164/438). Em geral, o gene mcr-1 foi encontrado em 90% (340/378) dos isolados de E. coli, com maiores prevalências encontradas em isola- dos de regiões litorâneas (96,5%, 191/198), humanos (95,6%, 111/116) e galinhas (91,8%, 123/134); para K. pneumoniae, o gene foi encontrado em 19,5% (32/164) dos isolados, com igual distribuição entre regiões e hospedeiros. Somente quatro isolados, dois de E. coli e dois de K. pneumoniae, demonstraram resistência fenotípica: o gene mcr-1 estava presente em ambas as cepas de E. coli, mas ausente nas de K. pneumoniae. Conclusões. Apesar da baixa prevalência de resistência fenotípica à colistina, a alta prevalência do gene mcr-1 em E. coli é preocupante. É preciso fiscalizar a proibição ao uso agropecuário de colistina no Equador e implementar o monitoramento contínuo da situação.

Molecular Tracking of the Origin of Vesicular Stomatitis Outbreaks in 2004 and 2018, Ecuador.

The Vesicular Stomatitis Virus (VSV) is an arbovirus causing vesicular stomatitis (VS) in livestock. There are two serotypes recognized: New Jersey (VSNJV) and Indiana (VSIV). The virus can be transmitted directly by contact or by vectors. In 2018, Ecuador experienced an outbreak of Vesicular Stomatitis (VS) in cattle, caused by VSNJV and VSVIV, with 399 cases reported distributed over 18 provinces. We determined the phylogenetic relationships among 67 strains. For the construction of phylogenetic trees, the viral phosphoprotein gene was sequenced, and trees were constructed based on the Maximum Likelihood method using 2004 outbreak strains from Ecuador (GenBank) and the 2018 sequences (this article). We built a haplotype network for VSNJV to trace the origin of the 2004 and 2018 epizootics through topology and mutation connections. These analyses suggest two different origins, one related to the 2004 outbreak and the other from a transmission source in 2018. Our analysis also suggests different transmission patterns; several small and independent outbreaks, most probably transmitted by vectors in the Amazon, and another outbreak caused by the movement of livestock in the Andean and Coastal regions. We recommend further research into vectors and vertebrate reservoirs in Ecuador to clarify the mechanisms of the reemergence of the virus.

Beta-Lactam Antibiotic Resistance Genes in the Microbiome of the Public Transport System of Quito, Ecuador.

Multidrug-resistant bacteria present resistance mechanisms against ß-lactam antibiotics, such as Extended-Spectrum Beta-lactamases (ESBL) and Metallo-ß-lactamases enzymes (MBLs) which are operon encoded in Gram-negative species. Likewise, Gram-positive bacteria have evolved other mechanisms through mec genes, which encode modified penicillin-binding proteins (PBP2). This study aimed to determine the presence and spread of ß-lactam antibiotic resistance genes and the microbiome circulating in Quito's Public Transport (QTP). A total of 29 station turnstiles were swabbed to extract the surface environmental DNA. PCRs were performed to detect the presence of 13 antibiotic resistance genes and to identify and to amplify 16S rDNA for barcoding, followed by clone analysis, Sanger sequencing, and BLAST search. ESBL genes blaTEM-1 and blaCTX-M-1 and MBL genes blaOXA-181 and mecA were detected along QPT stations, blaTEM being the most widely spread. Two subvariants were found for blaTEM-1, blaCTX-M-1, and blaOXA-181. Almost half of the circulating bacteria found at QPT stations were common human microbiota species, including those classified by the WHO as pathogens of critical and high-priority surveillance. ß-lactam antibiotic resistance genes are prevalent throughout QPT. This is the first report of blaOXA-181 in environmental samples in Ecuador. Moreover, we detected a new putative variant of this gene. Some commensal coagulase-negative bacteria may have a role as mecA resistance reservoirs.

Highin vitroactivity of gold and silver nanoparticles from Solanum mammosum L. against SARS-CoV-2 surrogate Phi6 and viral model PhiX174.

The search for new strategies to curb the spread of the SARS-CoV-2 coronavirus, which causes COVID-19, has become a global priority. Various nanomaterials have been proposed as ideal candidates to inactivate the virus; however, because of the high level of biosecurity required for their use, alternative models should be determined. This study aimed to compare the effects of two types of nanomaterials gold (AuNPs) and silver nanoparticles (AgNPs), recognized for their antiviral activity and affinity with the coronavirus spike protein using PhiX174 and enveloped Phi6 bacteriophages as models. To reduce the toxicity of nanoparticles, a species known for its intermediate antiviral activity,Solanum mammosumL. (Sm), was used. NPs prepared with sodium borohydride (NaBH4) functioned as the control. Antiviral activity against PhiX174 and Phi6 was analyzed using its seed, fruit, leaves, and essential oil; the leaves were the most effective on Phi6. Using the aqueous extract of the leaves, AuNPs-Sm of 5.34 ± 2.25 nm and AgNPs-Sm of 15.92 ± 8.03 nm, measured by transmission electron microscopy, were obtained. When comparing NPs with precursors, both gold(III) acetate and silver nitrate were more toxic than their respective NPs (99.99% at 1 mg ml-1). The AuNPs-Sm were less toxic, reaching 99.30% viral inactivation at 1 mg ml-1, unlike the AgNPs-Sm, which reached 99.94% at 0.01 mg ml-1. In addition, cell toxicity was tested in human adenocarcinoma alveolar basal epithelial cells (A549) and human foreskin fibroblasts. Gallic acid was the main component identified in the leaf extract using high performance liquid chromatography with diode array detection (HPLC-DAD). The FT-IR spectra showed the presence of a large proportion of polyphenolic compounds, and the antioxidant analysis confirmed the antiradical activity. The control NPs showed less antiviral activity than the AuNPs-Sm and AgNPs-Sm, which was statistically significant; this demonstrates that both theS. mammosumextract and its corresponding NPs have a greater antiviral effect on the surrogate Phi bacteriophage, which is an appropriate model for studying SARS-CoV-2.

Colistin resistance in Escherichia coli and Klebsiella pneumoniae in humans and backyard animals in Ecuador / Resistencia a la colistina de las bacterias Escherichia coli y Klebsiella pneumoniae en humanos y animales de granja en Ecuador / Resistência à colistina em Escherichia coli e Klebsiella pneumoniae em humanos e animais de quintal no Equador

ABSTRACT Objective. Colistin is an antibiotic of last resort for treating serious Gram-negative bacterial infections. However, the misuse of colistin, especially as an animal growth promoter, has contributed to increasing antimicrobial resistance, mediated mainly through plasmid transfer of the mcr-1 gene. This study assessed the prevalence of phenotypic and molecular colistin resistance in Escherichia coli and Klebsiella pneumoniae in Ecuador in healthy humans and their chickens and pigs. Methods. Fecal samples were collected from humans and their chickens and pigs in two rural coastal and Amazon regions between April and August 2020. Gram-negative bacteria were isolated and identified using conventional techniques. Phenotypic resistance was determined using the broth microdilution technique, and the mcr-1 gene was detected using conventional polymerase chain reaction. Results. A total of 438 fecal samples were obtained from 137 humans, 147 pigs and 154 chickens. The prevalence of E. coli isolates was 86.3% (378/438) and K. pneumoniae, 37.4% (164/438). Overall, the mcr-1 gene was found in 90% (340/378) of E. coli isolates, with higher prevalences found in isolates from coastal regions (96.5%, 191/198), humans (95.6%, 111/116) and chickens (91.8%, 123/134); for K. pneumoniae, the gene was found in 19.5% (32/164) of isolates, with equal distribution between regions and hosts. Only four isolates, two E. coli and two K. pneumoniae, showed phenotypic resistance: mcr-1 was present in both E. coli strains but absent in the K. pneumoniae strains. Conclusions. Despite a low prevalence of phenotypic resistance to colistin, the high prevalence of the mcr-1 gene in E. coli is of concern. Ecuador's ban on using colistin in animal husbandry must be enforced, and continual monitoring of the situation should be implemented.

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RESUMO Objetivo. A colistina é um antibiótico de último recurso para o tratamento de infecções graves por bactérias Gram-negativas. Entretanto, o uso indevido da colistina, principalmente como promotor de crescimento animal, tem contribuído para o aumento da resistência a antimicrobianos, principalmente por transferência horizontal do gene mcr-1 mediada por plasmídeos. Este estudo avaliou a prevalência de resistência fenotípica e molecular à colistina em Escherichia coli e Klebsiella pneumoniae no Equador em humanos hígidos e em galinhas e porcos por eles criados. Métodos. Entre abril e agosto de 2020, foram coletadas amostras de fezes de habitantes de duas regiões litorâneas e amazônicas do Equador e de galinhas e porcos por eles criados. Bactérias Gram-negativas foram isoladas e identificadas por meio de técnicas convencionais. A resistência fenotípica foi determinada pela técnica de microdiluição em caldo, e o gene mcr-1 foi detectado por reação em cadeia da polimerase convencional. Resultados. Foram obtidas 438 amostras fecais de 137 humanos, 147 suínos e 154 galinhas. A prevalência de isolados de E. coli foi de 86,3% (378/438), e de K. pneumoniae, 37,4% (164/438). Em geral, o gene mcr-1 foi encontrado em 90% (340/378) dos isolados de E. coli, com maiores prevalências encontradas em isolados de regiões litorâneas (96,5%, 191/198), humanos (95,6%, 111/116) e galinhas (91,8%, 123/134); para K. pneumoniae, o gene foi encontrado em 19,5% (32/164) dos isolados, com igual distribuição entre regiões e hospedeiros. Somente quatro isolados, dois de E. coli e dois de K. pneumoniae, demonstraram resistência fenotípica: o gene mcr-1 estava presente em ambas as cepas de E. coli, mas ausente nas de K. pneumoniae. Conclusões. Apesar da baixa prevalência de resistência fenotípica à colistina, a alta prevalência do gene mcr-1 em E. coli é preocupante. É preciso fiscalizar a proibição ao uso agropecuário de colistina no Equador e implementar o monitoramento contínuo da situação.

Whole-genome sequencing for surveillance of antimicrobial resistance in Ecuador: present and future implications / Secuenciación del genoma completo para la vigilancia de la resistencia a los antimicrobianos en Ecuador: implicaciones actuales y futuras / Sequenciamento do genoma completo para vigilância da resistência aos antimicrobianos no Equador: implicações presentes e futuras

ABSTRACT Whole-genome sequencing is becoming the gold standard for pathogen characterization and offers considerable advantages for understanding the evolution and dissemination of new determinants of antimicrobial resistance. Despite the benefits of whole-genome sequencing for pathogen characterization, implementation costs and lack of expertise may limit its use by public health laboratories. This article reviews the advantages of whole-genome sequencing for pathogen characterization and the current status of the use of whole-genome sequencing for antimicrobial resistance surveillance in Ecuador. A roadmap is suggested for including whole-genome sequencing for pathogen characterization based on the needs of the health reference institutions through alliances with Ecuadorian universities. Establishing a partnership between public health institutions and academia would be valuable for clinicians, policy-makers, and epidemiologists who could then take reasonable measures in those areas and establish a basis for adapting One Health strategies to tackle antimicrobial resistance in Ecuador.

RESUMEN La secuenciación del genoma completo, que está pasando a ser el estándar de referencia para la caracterización de agentes patógenos, ofrece ventajas considerables para comprender la evolución y la diseminación de los nuevos determinantes de la resistencia a los antimicrobianos. Sin embargo, a pesar de los beneficios que genera, los costos de ejecución y la falta de experiencia pueden limitar su uso por parte de los laboratorios de salud pública. En este artículo se evalúan las ventajas de la secuenciación del genoma completo para la caracterización de agentes patógenos y el estado actual del uso de la secuenciación del genoma completo en la vigilancia de la resistencia a los antimicrobianos en Ecuador. Se propone una hoja de ruta para incluir la secuenciación del genoma completo para la caracterización de agentes patógenos según las necesidades de las instituciones de salud de referencia, lo que se haría por medio de alianzas con universidades ecuatorianas. Establecer una asociación entre las instituciones de salud pública y los círculos académicos sería sumamente valioso para los médicos, los responsables de las políticas y los epidemiólogos, que podrían adoptar medidas razonables en sus ámbitos y sentar una base para adaptar las estrategias de "Una salud" a fin de abordar la resistencia a los antimicrobianos en Ecuador.

RESUMO O sequenciamento do genoma completo está se tornando o padrão ouro para a caracterização de patógenos e oferece vantagens consideráveis para a compreensão da evolução e disseminação de novos determinantes de resistência aos antimicrobianos. Apesar dos benefícios do sequenciamento do genoma completo para a caracterização de patógenos, os custos de implementação e a falta de especialização podem limitar seu uso pelos laboratórios de saúde pública. Este artigo analisa as vantagens do sequenciamento do genoma completo para a caracterização de patógenos e a situação atual do uso desta técnica para a vigilância da resistência aos antimicrobianos no Equador. Sugere-se um roteiro para incluir o sequenciamento de genomas completos para caracterização de patógenos com base nas necessidades das instituições de saúde de referência, por meio de alianças com universidades equatorianas. A criação de uma parceria entre instituições de saúde pública e entidades acadêmicas seria valiosa para clínicos, formuladores de políticas e epidemiologistas, que poderiam, assim, tomar medidas razoáveis nessas áreas e estabelecer uma base para adaptar estratégias de Saúde Única para combater a resistência aos antimicrobianos no Equador.

High Prevalence of Prototheca bovis Infection in Dairy Cattle with Chronic Mastitis in Ecuador.

The genus Prototheca, a unicellular, non-photosynthetic, yeast-like microalgae, is a pathogen of concern for the dairy industry. It causes bovine mastitis that currently cannot be cured, and hence generates significant economic losses in milk production. In this study, for the first time in Ecuador, we identify Prototheca bovis as the etiologic agent of chronic mastitis in dairy cattle. Milk samples (n = 458) of cows with chronic mastitis were cultured on Sabouraud Dextrose Agar (SDA). Microscopy and cytB gene sequencing were used to identify Prototheca, whereby Prototheca bovis was isolated from 15.1% (n = 69) of the milk samples, one of the highest infection rates that can be found in the literature in a "non-outbreak" situation. No other Prototheca species were found. We were unable to isolate the alga from environmental samples. We showed that P. bovis was relatively resistant to disinfectants used to sterilize milking equipment on the cattle farms where it was isolated. We discuss how to avoid future infection and also hypothesize that the real prevalence of Prototheca infection in bovine mastitis is probably much higher than what was detected. We recommend a protocol to increase the diagnostic yield in the bacteriology laboratory.

The Re-Identification of Previously Unidentifiable Clinical Non-Tuberculous Mycobacterial Isolates Shows Great Species Diversity and the Presence of Other Acid-Fast Genera.

Non-tuberculous mycobacteria that cannot be identified at the species level represent a challenge for clinical laboratories, as proper species assignment is key to implementing successful treatments or epidemiological studies. We re-identified forty-eight isolates of Ziehl-Neelsen (ZN)-staining-positive "acid-fast bacilli" (AFB), which were isolated in a clinical laboratory and previously identified as Mycobacterium species but were unidentifiable at the species level with the hsp65 PCR restriction fragment length polymorphism analysis (PRA). As most isolates also could not be identified confidently via 16S, hsp65, or rpoB DNA sequencing and a nBLAST search analysis, we employed a phylogenetic method for their identification using the sequences of the 16S rDNA, which resulted in the identification of most AFB and a Mycobacterium species diversity not found before in our laboratory. Most were rare species with only a few clinical reports. Moreover, although selected with the ZN staining as AFB, not all isolates belonged to the genus Mycobacterium, and we report for the first time in Latin America the isolation of Nocardia puris, Tsukamurella pulmosis, and Gordonia sputi from sputum samples of symptomatic patients. We conclude that ZN staining does not differentiate between the genus Mycobacterium and other genera of AFB. Moreover, there is a need for a simple and more accurate tree-based identification method for mycobacterial species. For this purpose, and in development in our lab, is a web-based identification system using a phylogenetic analysis (including all AFB genera) based on 16S rDNA sequences (and in the future multigene datasets) and the closest relatives.

Extended-Spectrum Beta-Lactamases Producing Escherichia coli in South America: A Systematic Review with a One Health Perspective.

Purpose: Extended-spectrum beta-lactamase-producing (ESBL) Enterobacteriaceae, which includes Escherichia coli, has emerged as a global health threat. ESBL enzymes including CTX-M, TEM, and SHV are the most detected. Here, a systematic review was developed to assess the status of ESBLs in E. coli considering studies performed in the human, animal, food, and environmental realms in South America. Methods: Following PRISMA guidelines, a systematic review was performed using the PubMed database as a primary source to identify studies containing data on ESBL-producing E. coli in South America. To obtain a comprehensive sample, studies in English, Spanish, and Portuguese were included from 1990 to April 2021. Inclusion such as the reporting of sample origin and diagnostic method and exclusion criteria such as review/letter articles were established to complete data extraction steps. Results: Amongst 506 articles retrieved, 130 met the inclusion criteria. Brazil reported 65 (50%) of publications, followed by Argentina, and Ecuador with 11.5% each. According to the category of studies, human studies represented the 56%, animals the 20%, environmental the 11%, and food studies the 6%. Interestingly, studies assessing more than one category (ie, interdisciplinary) represented the 7%. Prevalence of ESBL producing E. coli in animal, food, and environmental studies was widely superior compared to human sources. In clinical studies, Brazil presented the greatest diversity in terms of ESBLs, featuring CTX-M, TEM, SHV, TOHO, OXA, and AmpC. CTX-M enzymes were the most frequent variants with 89.4% detections. Conclusion: The present One Health review of 130 studies conducted over the past 21 years found ESBLs producing E. coli distributed across human, animal, food, and environmental samples across South America. There is a need to increment studies in underrepresented countries and to strengthen multi-sectoral antimicrobial resistance research and surveillance. This information can be used as basis for subsequent implementation of monitoring programs, targeting potential critical points of transmission sources.

Clonality and Persistence of Multiresistant Methicillin-Resistant Coagulase-Negative Staphylococci Isolated from the Staff of a University Veterinary Hospital.

The aim of this study was to characterize methicillin-resistant coagulase-negative staphylococci (MRCoNS) isolates from the healthy staff of a university veterinary hospital in order to assess their importance as a reservoir of antimicrobial resistance and to determine their population structure and evolution. The study duration was over two years (2020-2021), 94 individuals were analyzed in duplicate, and 78 strains were obtained. The overall prevalence of methicillin-resistant strains detected throughout the study was 61.7%, with point prevalence values of 53.2% in 2020 and 31.5% in 2021. A total of 19.1% of the individuals analyzed were carriers throughout the study. The most frequently identified MRCoNs were Staphylococcus epidermidis (92.3%) and S. warneri (3.8%). A total of 75.6% of the isolates obtained showed the development of multi-resistance, preferentially against erythromycin, gentamicin, and tetracycline, and to a lesser extent against fusidic acid, norfloxacin, and clindamycin; these antimicrobials are frequently used in the veterinary field. Although most of the S. epidermidis isolates obtained showed wide genetic variability and low dispersion, which are characteristic of community-associated isolates, a small number of strains spread between individuals in close physical proximity and were maintained over time, forming stable clones. These clones generally maintained the same type of staphylococcal cassette chromosome mec (SCCmec) and had a similar antimicrobial resistance pattern.