SARS-CoV-2 infection in brown-headed spider monkeys (Ateles fusciceps) at a wildlife rescue center on the coast of Ecuador-South America.

Human populations can be affected in unpredictable ways by the emergence and spread of zoonotic diseases. The COVID-19 (coronavirus disease of 2019) pandemic was a reminder of how devastating these events can be if left unchecked. However, once they have spread globally, the impact of these diseases when entering non-exposed wildlife populations is unknown. The current study reports the infection of brown-headed spider monkeys (Ateles fusciceps) at a wildlife rescue center in Ecuador. Four monkeys were hospitalized, and all tested positive for SARS-CoV-2 (Severe acute respiratory syndrome coronavirus 2) by RT-qPCR (Quantitative Reverse Transcription PCR). Fecal samples (n = 12) from monkeys at the rescue center also tested positive; three zookeepers responsible for feeding and deworming the monkeys also tested positive, suggesting human-animal transmission. Whole genome sequencing identified most samples' omicron clade 22B BA.5 lineage. These findings highlight the threat posed by an emerging zoonotic disease in wildlife species and the importance of preventing spillover and spillback events during epidemic or pandemic events.IMPORTANCEAlthough COVID-19 (coronavirus disease of 2019) has been primarily contained in humans through widespread vaccination, the impact and incidence of SARS-CoV-2 (Severe acute respiratory syndrome coronavirus) and its transmission and epidemiology in wildlife may need to be addressed. In some natural environments, the proximity of animals to humans is difficult to control, creating perfect scenarios where susceptible wildlife can acquire the virus from humans. In these places, it is essential to understand how transmission can occur and to develop protocols to prevent infection. This study reports the infection of brown-headed spider monkeys with SARS-CoV-2, a red-listed monkey species, at a wildlife recovery center in Ecuador. This study reports the infection of brown-headed spider monkeys with SARS-CoV-2, indicating the potential for transmission between humans and wildlife primates and the importance of preventing such events in the future.

Endogenous origin of Pseudomonas aeruginosa infecting hospitalized patients in Ecuador.

Recent evidence suggests that Pseudomonas aeruginosa, a bacterium that has the ability to cause deadly infections in hospitalized patients, could originate in the patient's own flora. We employed the Oxford Nanopore platform to obtain whole genome sequences (WGS) from clinical and rectal screen P. aeruginosa strains belonging to 15 patients from two hospitals. Our study found evidence that clinical and rectal isolates were clonal, with some evidence suggesting that the infecting strain was present in the patient's intestine at the time of admission, ruling out hospital acquisition. The use of WGS analysis is crucial to detect alternative sources of P. aeruginosa to develop new preventive measures against these serious infections.

IS26 drives the dissemination of bla CTX-M genes in an Ecuadorian community.

IMPORTANCE: The horizontal gene transfer events are the major contributors to the current spread of CTX-M-encoding genes, the most common extended-spectrum ß-lactamase (ESBL), and many clinically crucial antimicrobial resistance (AMR) genes. This study presents evidence of the critical role of IS26 transposable element for the mobility of bla CTX-M gene among Escherichia coli isolates from children and domestic animals in the community. We suggest that the nucleotide sequences of IS26-bla CTX-M could be used to study bla CTX-M transmission between humans, domestic animals, and the environment, because understanding of the dissemination patterns of AMR genes is critical to implement effective measures to slow down the dissemination of these clinically important genes.

Distribution of blaCTX-M-gene variants in E. coli from different origins in Ecuador.

The increasing abundance of extended spectrum (ß-lactamase (ESBL) genes in E. coli, and other commensal and pathogenic bacteria, endangers the utility of third or more recent generation cephalosporins, which are major tools for fighting deadly infections. The role of domestic animals in the transmission of ESBL carrying bacteria has been recognized, especially in low- and middle-income countries, however the horizontal gene transfer of these genes is difficult to assess. Here we investigate blaCTX-M gene diversity (and flanking nucleotide sequences) in E. coli from chicken and humans, in an Ecuadorian rural community and from chickens in another location in Ecuador. The blaCTX-M associated sequences in isolates from humans and chickens in the same remote community showed greater similarity than those found in E. coli in a chicken industrial operation 200 km away. Our study may provide evidence of blaCTX-M transfer between chickens and humans in the community.

Delayed acquisition of airway commensals in antibiotic naïve children and its relationship with wheezing in rural Ecuador.

Introduction: The hygiene hypothesis identified a relationship between living in rural areas and acquiring protective environmental factors against the development of asthma and atopy. In our previous study, we found a correlation between particular bacterial species and early-onset wheezing in infants from the rural tropics of Ecuador who were corticosteroid-naïve and had limited antibiotic exposure. We now describe a longitudinal study of infants conducted to determine the age-related changes of the microbiome and its relationship with wheezing. Methods: We performed an amplicon sequencing of the 16S rRNA bacterial gene from the oropharyngeal samples obtained from 110 infants who had a history of recurrent episodic wheezing sampled at different ages (7, 12, and 24 months) and compared it to the sequencing of the oropharyngeal samples from 150 healthy infants sampled at the same time points. Bioinformatic analyses were conducted using QIIME and R. Results: As expected, the microbiota diversity consistently increased as the infants grew older. Considering age-based microbiota changes, we found that infants with wheeze had significantly lower species richness than the healthy infants at 7 months, but not at 12 or 24 months. Most of the core and accessory organisms increased in abundance and prevalence with age, except for a few which decreased. At 7 months of age, infants with wheeze had notably higher levels of a single Streptococcus operational taxonomic unit and core microbiota member than controls. Conclusions: In a cohort with limited antibiotic and corticosteroid use, a progressively more complex and diverse respiratory microbial community develops with age. The respiratory microbiota in early life is altered in infants with wheeze, but this does not hold true in older infants.

Multi-level analysis of the gut-brain axis shows autism spectrum disorder-associated molecular and microbial profiles.

Autism spectrum disorder (ASD) is a neurodevelopmental disorder characterized by heterogeneous cognitive, behavioral and communication impairments. Disruption of the gut-brain axis (GBA) has been implicated in ASD although with limited reproducibility across studies. In this study, we developed a Bayesian differential ranking algorithm to identify ASD-associated molecular and taxa profiles across 10 cross-sectional microbiome datasets and 15 other datasets, including dietary patterns, metabolomics, cytokine profiles and human brain gene expression profiles. We found a functional architecture along the GBA that correlates with heterogeneity of ASD phenotypes, and it is characterized by ASD-associated amino acid, carbohydrate and lipid profiles predominantly encoded by microbial species in the genera Prevotella, Bifidobacterium, Desulfovibrio and Bacteroides and correlates with brain gene expression changes, restrictive dietary patterns and pro-inflammatory cytokine profiles. The functional architecture revealed in age-matched and sex-matched cohorts is not present in sibling-matched cohorts. We also show a strong association between temporal changes in microbiome composition and ASD phenotypes. In summary, we propose a framework to leverage multi-omic datasets from well-defined cohorts and investigate how the GBA influences ASD.

Evaluation of changes in the faecal resistome associated with children's exposure to domestic animals and food animal production.

OBJECTIVES: The paediatric gut microbiota is a reservoir of antimicrobial resistance genes. Environmental factors such as a child's exposure to faecal contamination and antimicrobial resistance genes of animal origin likely shape the resistome of infants and children. This study measured how different levels of exposure to domestic or food animals affect the structure of the intestinal resistome in children between 1 and 7 years of age. METHODS: One hundred nineteen faecal samples from 39 children were analysed according to the level of exposure to domestic or food animals and categorized into three risk groups. Using high-throughput sequencing with an Illumina NovaSeq 6000 SP platform, we performed faecal resistome analyses using the ResFinder database. Additionally, ResistoXplorer was used to characterize the resistomes of children differentially exposed to domestic animals. RESULTS: Our data indicated that specific antimicrobial resistance genes such as those that confer resistance to MATFPR (macrolide, aminoglycoside, tetracycline, fluoroquinolone, phenicol, and rifamycin) and tetracyclines were statistically less abundant in the group of children without exposure to animals (group 2), compared with the groups exposed to domestic and food animals (groups 1 and 3). However, the overall resistome structure among the children was not affected by the different levels of exposure to animals. CONCLUSIONS: This study suggests that animal exposure is a risk factor for young children acquiring specific antimicrobial resistance genes from domestic animals or animal production areas. However, the overall resistome structure was not affected.

Occurrence of SARS-CoV-2 reinfections at regular intervals in Ecuador.

SARS-CoV-2 reinfection is defined as a new infection with a different virus variant in an individual who has already recovered from a previous episode of COVID-19. The first case of reinfection in the world was described in August 2020, since then, reinfections have increased over time and their incidence has fluctuated with specific SARS-CoV-2 variant waves. Initially, reinfections were estimated to represent less than 1% of total COVID-19 infections. With the advent of the Omicron variant, reinfections became more frequent, representing up to 10% of cases (based on data from developed countries). The frequency of reinfections in Latin America has been scarcely reported. The current study shows that in Ecuador, the frequency of reinfections has increased 10-fold following the introduction of Omicron, after 22 months of surveillance in a single center of COVID-19 diagnostics. Suspected reinfections were identified retrospectively from a database of RT-qPCR-positive patients. Cases were confirmed by sequencing viral genomes from the first and second infections using the ONT MinION platform. Monthly surveillance showed that the main incidence peaks of reinfections were reached within four to five months, coinciding with the increase of COVID-19 cases in the country, suggesting that the emergence of reinfections is related to higher exposure to the virus during outbreaks. This study performed the longest monitoring of SARS-CoV-2 reinfections, showing an occurrence at regular intervals of 4-5 months and confirming a greater propensity of Omicron to cause reinfections.

Corrigendum: Extended-Spectrum Beta-Lactamase Producing-Escherichia coli Isolated From Irrigation Waters and Produce in Ecuador.

[This corrects the article DOI: 10.3389/fmicb.2021.709418.].

The impact of genetic recombination on pathogenic Leptospira.

Leptospirosis is the most common zoonosis worldwide, and is increasingly common in poor urban communities, where there is inadequate sewage disposal and abundance of domestic and peridomestic animals. There are many risk factors associated with the disease, such as contaminated water exposure, close contact with animals, floods, recreational activities related to water, wet agriculture. The symptoms of leptospirosis are common to other infectious diseases and, if not treated, it can lead to meningitis, liver failure, kidney damage and death. Leptospirosis is caused by 38 pathogenic species of Leptospira, which are divided in almost 30 serogroups and more than 300 serovars. The serological classification (serogroups and serovars) is based on the expression of distinct lipopolysaccharide (LPS) antigens. These antigens are also associated to protective immunity; antibodies against a serovar protect from any member of the same serovar. Serologic and phylogenetic analyses are not congruent probably due to genetic recombination of LPS genes among different leptospiral species. To analyze the importance of recombination in leptospiral evolution, we performed a gene-by-gene tree topology comparison on closed genomes available in public databases at two levels: among core genomes of pathogenic species (34 recombinant among 1213 core genes), and among core genomes of L. interrogans isolates (178/798). We found that most recombinant genes code for proteins involved in translation, ribosomal structure and biogenesis, but also for cell wall, membrane and envelope biogenesis. Besides, our final results showed that half of LPS genes are recombinant (18/36). This is relevant because serovar classification and vaccine development are based on these epitopes. The frequent recombination of LPS-associated genes suggests that natural selection is promoting the survival of recombinant lineages. These results may help understanding the factors that make Leptospira a successful pathogen.

A longitudinal study of dominant E. coli lineages and antimicrobial resistance in the gut of children living in an upper middle-income country.

OBJECTIVES: The gastrointestinal tract constitutes a complex and diverse ecosystem. Escherichia coli is one of the most frequently studied and characterised species in the gut ecosystem; nevertheless, there has been little research to determine their diversity and population dynamics in the intestines of children over time. We analysed the turnover or dominant E. coli isolates in children faecal matter during 1 year. METHODS: In this prospective study, a fresh faecal sample was obtained from children longitudinally over one year (30 faecal samples at sampling period 1 and 22 faecal samples at sampling periods 2 and 3). From each stool sample, five E. coli colonies were randomly selected (n = 405 E. coli isolates total) in order to characterize the genotype and phenotypic antimicrobial resistance patterns. RESULTS: We were unable to find same E. coli dominant clone in faecal matter from 30 children in different sampling periods. Whole-genome sequencing of three isolates belonging to ST131 found in one child during the sampling period I and II indicated that isolates were three different ST 131 clones that carried extended-spectrum ß-lactamase (ESBL) genes. CONCLUSION: We found that all numerically dominant E. coli lineages in children's intestines were transient colonisers, and antimicrobial resistance phenotypes of these strains varied significantly over time without any apparent selective force.

Characterization of the Immune Resistance of Severe Acute Respiratory Syndrome Coronavirus 2 Mu Variant and the Robust Immunity Induced by Mu Infection.

BACKGROUND: We have recently revealed that the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Mu variant shows a pronounced resistance to antibodies elicited by natural SARS-CoV-2 infection and vaccination. METHODS: However, it remains unclear which mutations determine the resistance of SARS-CoV-2 Mu to antiviral sera. In addition, it is unclear how SARS-CoV-2 Mu infection induces antiviral immunity. RESULTS: In this study, we reveal that the 2 mutations in the SARS-CoV-2 Mu spike protein, YY144-145TSN and E484K, are responsible for the resistance to coronavirus disease 2019 convalescent sera during early 2020 and vaccine sera. CONCLUSIONS: It is notable that the convalescent sera of SARS-CoV-2 Mu-infected individuals are broadly antiviral against Mu as well as other SARS-CoV-2 variants of concern and interest.

Dynamics of Microbial Communities during the Removal of Copper and Zinc in a Sulfate-Reducing Bioreactor with a Limestone Pre-Column System.

Biological treatment using sulfate-reducing bacteria (SRB) is a promising approach to remediate acid rock drainage (ARD). Our purpose was to assess the performance of a sequential system consisting of a limestone bed filter followed by a sulfate-reducing bioreactor treating synthetic ARD for 375 days and to evaluate changes in microbial composition. The treatment system was effective in increasing the pH of the ARD from 2.7 to 7.5 and removed total Cu(II) and Zn(II) concentrations by up to 99.8% and 99.9%, respectively. The presence of sulfate in ARD promoted sulfidogenesis and changed the diversity and structure of the microbial communities. Methansarcina spp. was the most abundant amplicon sequence variant (ASV); however, methane production was not detected. Biodiversity indexes decreased over time with the bioreactor operation, whereas SRB abundance remained stable. Desulfobacteraceae, Desulfocurvus, Desulfobulbaceae and Desulfovibrio became more abundant, while Desulfuromonadales, Desulfotomaculum and Desulfobacca decreased. Geobacter and Syntrophobacter were enriched with bioreactor operation time. At the beginning, ASVs with relative abundance <2% represented 65% of the microbial community and 21% at the end of the study period. Thus, the results show that the microbial community gradually lost diversity while the treatment system was highly efficient in remediating ARD.

Saccharomyces boulardii CNCM I-745 supplementation modifies the fecal resistome during Helicobacter pylori eradication therapy.

BACKGROUND: The gut microbiota is a significant reservoir of antimicrobial resistance genes (ARGs). The use and misuse of antimicrobials can select multi-resistant bacteria and modify the repertoire of ARGs in the gut. Developing effective interventions to manipulate the intestinal resistome would allow us to modify the antimicrobial resistance risk. MATERIALS AND METHODS: Applying shotgun metagenomics, we compared the composition of fecal resistome from individuals treated with triple therapy for Helicobacter pylori plus Saccharomyces boulardii CNCM-I 745 (Sb) versus triple antibiotherapy without S. boulardii (control) before, after, and one month after treatments. DNA samples were sequenced on an Illumina NovaSeq 6000 platform. Reads were trimmed and filtered for quality, and the reads classified as host genome were removed from further analysis. We used the ResFinder database for resistome analysis and the web-based tool ResistoXplorer and RStudio for graphical representation and statistical analysis. RESULTS: We identified 641 unique ARGs in all fecal samples, conferring resistance to 18 classes of antibiotics. The most prevalent ARGs found in at least 90% of the samples before the treatments were against tetracyclines, MLS-B (macrolide, lincosamide, and streptogramin B), beta-lactams, and aminoglycosides. Differential abundance analysis allowed the identification of ARGs significantly different between treatment groups. Thus, immediately after the treatments, the abundance of ARGs that confer resistance to lincosamides, tetracyclines, MLS-B, and two genes in the beta-lactam class (cfxA2 and cfxA3) was significantly lower in the group that received Sb than in the control group (edgeR, FDR <0.05). CONCLUSION: Our study demonstrated that the addition of S. boulardii CNCM-I 745 to the conventional antibiotic eradication therapy for H. pylori reduced the abundance of ARGs, particularly those genes that confer resistance to lincosamides, tetracyclines, MLS-B, and a few genes in the beta-lactams class.

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

Extended-spectrum ß-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.

Extended-Spectrum Beta-Lactamase Producing-Escherichia coli Isolated From Irrigation Waters and Produce in Ecuador.

In cities across the globe, the majority of wastewater - that includes drug resistant and pathogenic bacteria among other contaminants - is released into streams untreated. This water is often subsequently used for irrigation of pastures and produce. This use of wastewater-contaminated streams allows antibiotic-resistant bacteria to potentially cycle back to humans through agricultural products. In this study, we investigated the prevalence of extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli isolated from produce and irrigation water across 17 provinces of Ecuador. A total of 117 vegetable samples, 119 fruit samples, and 38 irrigation water samples were analyzed. Results showed that 11% of the samples were positive for E. coli including 11 irrigation water samples (29%), and samples of 13 vegetables (11%), and 11 fruits (9%). Among the 165 E. coli isolates cultured, 96 (58%) had the ESBL phenotype, and 58% of ESBL producing E. coli came from irrigation water samples, 11% from vegetables, and 30% from fruits. The bla CTX-M - 55, bla CTX-M 65, and bla CTX-M 15 genes were the most frequently found gene associated with the ESBL phenotype and coincided with the bla CTX-M alleles associated with human infections in Ecuador. Three isolates had the mcr-1 gene which is responsible for colistin resistance. This report provides evidence of the potential role of irrigation water in the growing antimicrobial resistance crisis in Ecuador.

Genomic epidemiology of SARS-CoV-2 transmission lineages in Ecuador.

Characterisation of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) genetic diversity through space and time can reveal trends in virus importation and domestic circulation and permit the exploration of questions regarding the early transmission dynamics. Here, we present a detailed description of SARS-CoV-2 genomic epidemiology in Ecuador, one of the hardest hit countries during the early stages of the coronavirus-19 pandemic. We generated and analysed 160 whole genome sequences sampled from all provinces of Ecuador in 2020. Molecular clock and phylogeographic analysis of these sequences in the context of global SARS-CoV-2 diversity enable us to identify and characterise individual transmission lineages within Ecuador, explore their spatiotemporal distributions, and consider their introduction and domestic circulation. Our results reveal a pattern of multiple international importations across the country, with apparent differences between key provinces. Transmission lineages were mostly introduced before the implementation of non-pharmaceutical interventions, with differential degrees of persistence and national dissemination.

SARS-CoV-2 detection and sequencing in heart tissue associated with myocarditis and persistent arrhythmia: A case report.

BACKGROUND: SARS-CoV-2 uses the human cell receptor angiotensin-converting enzyme (ACE2). ACE2 is widely present in the cardiovascular system including the myocardium and the conduction system. COVID-19 patients that present severe symptoms have been reported to have complications involving myocardial injuries caused by the virus. Here we report the detection of SARS-CoV-2 by whole genome sequencing in the endocardium of a patient with severe bradycardia. CASE PRESENTATION: We report a case of a 34-year-old male patient with COVID-19 tested by PCR, he started with gastrointestinal symptoms, however, he quickly deteriorated his hemodynamic state by means of myocarditis and bradycardia. After performing an endocardium biopsy, it was possible to identify the presence of SARS-CoV-2 in the heart tissue and to sequence its whole genome using the ARTIC-Network protocol and a modified tissue RNA extraction method. The patient's outcome was improved after a permanent pacemaker was implanted. CONCLUSIONS: It was possible to identify a SARS-CoV-2 clade 20A in the endocardium of the reported patient.

SARS-CoV-2 genome sequencing from COVID-19 in Ecuadorian patients: a whole country analysis.

SARS-CoV-2, the etiological agent of COVID-19, was first described in Wuhan, China in December 2019 and has now spread globally. Ecuador was the second country in South America to confirm cases and Guayaquil was one of the first cities in the world to experience high mortality due to COVID-19. The aim of this study was to describe the lineages circulating throughout the country and to compare the mutations in local variants, to the reference strain. In this work we used the MinION platform (Oxford Nanopore Technologies) to sequence the whole SARS-CoV-2 genomes of 119 patients from all provinces of Ecuador, using the ARTIC network protocols. Our data from lineage assignment of the one hundred and nineteen whole genomes revealed twenty different lineages. All genomes presented differences in the S gene compared to the Wuhan reference strain, being the D614G amino acid replacement the most common change. The B.1.1.119 lineage was the most frequent and was found in several locations in the Coast and Andean region. Three sequences were assigned to the new B.1.1.7 lineage. Our work is an important contribution to the understanding of the epidemiology of SARS-CoV-2 in Ecuador and South America.