Isolates Of Human Origin Research Articles

Exploring AMR and virulence in Klebsiella pneumoniae isolated from humans and pet animals: A complement of phenotype by WGS-derived profiles in a One Health study in Egypt

Klebsiella pneumoniae is a ubiquitous nosocomial pathogen associated with various types of infections in hospitalized patients and different animal species. In the current study, 49 Klebsiella strains isolated from humans, dogs, and cats were investigated using NGS technology. MALDI-TOF failed to identify newly discovered K. variicola and K. quasipneumoniae isolates correctly. MLST analysis revealed different sequence types among K. pneumoniae isolates, and the most frequent STs were ST29, ST219, and ST37. Three ST23 that are generally known as hypervirulent type were identified but they lacked major discriminatory determinants for hypervirulent K. pneumoniae (hvKp). K. pneumoniae isolates showed high diversity, and several isolates from humans and animals were assigned to the same ST and were almost identical. Isolates from humans exhibited more pronounced resistance patterns compared to the animal isolates. High levels of resistance were observed for piperacillin, trimethoprim/sulfamethoxazole, and cephalosporins, and resistance to carbapenem compounds was only found in isolates of human origin. Three strains of human origin were extensively drug-resistant (XDR). A diverse range of resistance genes primarily confer resistance to beta-lactams., phenicol/quinolone, aminoglycoside, macrolide, sulfonamides, and fosfomycin were identified in silico. However, there were inconsistencies between the phenotypic characterization of isolates and the set of resistance genes detected in silico in this set of Klebsiella isolates. Further research using a larger number of isolates from various sources is necessary to fully comprehend the relationship between the presence of antimicrobial resistance determinants and phenotypic data. It is also necessary to monitor the spread of K. pneumoniae from a One Health perspective in Egypt.

Analysis of twelve genomes of the bacterium Kerstersia gyiorum from brown-throated sloths (Bradypus variegatus), the first from a non-human host.

Kerstersia gyiorum is a Gram-negative bacterium found in various animals, including humans, where it has been associated with various infections. Knowledge of the basic biology of K. gyiorum is essential to understand the evolutionary strategies of niche adaptation and how this organism contributes to infectious diseases; however, genomic data about K. gyiorum is very limited, especially from non-human hosts. In this work, we sequenced 12 K. gyiorum genomes isolated from healthy free-living brown-throated sloths (Bradypus variegatus) in the Parque Estadual das Fontes do Ipiranga (São Paulo, Brazil), and compared them with genomes from isolates of human origin, in order to gain insights into genomic diversity, phylogeny, and host specialization of this species. Phylogenetic analysis revealed that these K. gyiorum strains are structured according to host. Despite the fact that sloth isolates were sampled from a single geographic location, the intra-sloth K. gyiorum diversity was divided into three clusters, with differences of more than 1,000 single nucleotide polymorphisms between them, suggesting the circulation of various K. gyiorum lineages in sloths. Genes involved in mobilome and defense mechanisms against mobile genetic elements were the main source of gene content variation between isolates from different hosts. Sloth-specific K.gyiorum genome features include an IncN2 plasmid, a phage sequence, and a CRISPR-Cas system. The broad diversity of defense elements in K. gyiorum (14 systems) may prevent further mobile element flow and explain the low amount of mobile genetic elements in K. gyiorum genomes. Gene content variation may be important for the adaptation of K. gyiorum to different host niches. This study furthers our understanding of diversity, host adaptation, and evolution of K. gyiorum, by presenting and analyzing the first genomes of non-human isolates.

Biofilm Production Ability of Streptococcus dysgalactiae Subsp. equisimilis: Associations with Host Species, Lancefield Group, Source, Clonal Complex, and Virulence-Associated Genes.

We assessed the biofilm production ability (BPA) of noninvasive Streptococcus dysgalactiae subsp. equisimilis (SDSE) in humans and companion animals and determined the relationship between bacterial populations with BPA and other host and microbiological features. Sixty-four isolates from companion animals and humans were collected along with host information. We measured BPA using crystal violet staining, in addition to emm typing, multilocus sequence typing, antimicrobial resistance (AMR) phenotyping/genotyping, and virulence-associated gene (VAG) detecting (prtF1-prtF2-lmb-cbp-sicG-srtp1-srtp2-brpA). Differences in the BPA of SDSE from different hosts and sources and different Lancefield groups were assessed. We analyzed the associations between populations with and without BPA (strong, moderate, weak, and no biofilm producers) and emm types, sequence types/clonal complexes (CCs), AMR phenotypes/genotypes, and VAG types. Seventeen, twenty-four, and twelve isolates were strong, moderate, and weak biofilm producers, respectively; eleven showed no BPA. There was a difference in the distribution of populations with BPA between human and animal origins and between isolates of groups G and C. We found an association between populations with BPA and the eye and ear source (vs. the pus and skin source). A relationship was observed between the populations with BPA and CC127 (vs. CC17). We observed no association between the populations with BPA and AMR phenotype/genotype. There was an association between the distribution of populations with BPA and srtp1 expression. Our observations suggest potential associations between populations with BPA and the host species, Lancefield group, source, CC, and VAG type.

First record of the human infection of Brucella melitensis in Kyrgyzstan: evidence from whole-genome sequencing-based analysis.

Brucellosis, a zoonosis mainly transmitted by consumption of unpasteurized dairy products as well as direct contact with infected animals, is endemic in Kyrgyzstan. However, Brucella species in humans have not been investigated and the origin of the disease remains poorly known in wide parts of Сentral Asia. Thus, molecular characterization of the circulating strains is a critical first step in understanding Brucella diversity in the country. In this study, isolates were collected from patients with suspected brucellosis from different regions in Kyrgyzstan between 2019 and 2020. The detection and identification of Brucella was carried out by Bruce-ladder PCR. Next generation sequencing was used to sequence the 89 Brucella isolates, which were genotyped by cgSNP and cgMLST to identify epidemiological connection between Brucella isolates as well as placing them in the context of the global Brucella phylogeny. The Brucella strains isolated from all regions of Kyrgyzstan were identified as B. melitensis. Based on cgSNP analysis, 18 sequence types were differentiated. The highest numbers of different sequence types were found in Batken (n = 8), Osh (n = 8) and Jalal-Abad (n = 6) oblasts. According to cgSNP and cgMLST analyses, different B. melitensis lineages circulate in Kyrgyzstan, all of them belonging to the Eastern Mediterranean group of the global Brucella phylogeny with the highest similarity to strains from Turkmenistan, Iran and Turkey. In the present study, B. melitensis was identified as a causative agent of human brucellosis in Kyrgyzstan and different lineages could be identified. Since this study focused on isolates of human origin, the identity of Brucella species and lineages circulating among animal populations remains elusive. Implementing culture techniques and use of most recent molecular, bioinformatic and epidemiological tools are needed to set up a One Health approach to combat brucellosis in Kyrgyzstan. Further, other Сentral Asian countries need to take part in this effort as brucellosis is a transboundary disease in these regions.

Withdrawal of antibiotic growth promoters in China and its impact on the foodborne pathogen Campylobacter coli of swine origin.

Antibiotic growth promoters (AGPs) have been used as feed additives to improve feed efficiency in food animals for more than six decades. However, the wide use of AGPs has led to the emergence of antibiotic-resistant pathogens of animal origin, posing a significant threat to food safety and public health. China prohibited the addition of AGPs to animal feed from July 2020. The impacts caused by the withdrawal of AGPs on the prevalence and antibiotic resistance of foodborne pathogens have not been illustrated. Here, a total of 471 strains of Campylobacter were isolated from pigs from three pig farms and two slaughterhouses in Sichuan Province for 4 consecutive years (2018–2021), including 2 years before and 2 years after the ban on AGPs in China. The isolation rate of Campylobacter had a slight increase after prohibiting the addition of AGPs to the feed. Contrary to what we expected, the antibiotic susceptibility test and WGS data showed that the antibiotic resistance to gentamicin and florfenicol and the abundance of virulence genes increased significantly after the ban of AGPs. Comparison of the isolates of swine origin with isolates of human origin indicated the potential of antibiotic-resistant Campylobacter transmission from pigs to humans. These data suggested that phasing out AGPs may lead to increased use of therapeutic antimicrobials, promoting the prevalence and transmission of both antibiotic resistance and virulence genes.

Evaluation of tolerance to artificial gastroenteric juice and fermentation characteristics of Lactobacillus strains isolated from human.

Fifty‐seven strains of Lactobacillus were isolated from fecal samples of healthy young people in Tibet, Xinjiang, and Inner Mongolia using pure culture methods. Lactobacillus ruminis and Lactobacillus gasseri were the dominant Lactobacillus species isolated from the intestinal microflora, accounting for 54.4% and 14.0% of the total isolates, respectively. Isolated strains were identified by 16S rRNA sequencing, and their tolerance to gastric acid and bile salt, and fermentation characteristics were evaluated. The results of experiments in vitro showed that nine of the isolated strains of Lactobacillus grew well at pH 3.0. After 11 h of incubation in artificial digestive juices, the isolated L. plantarum and the control strain L. plantarum P8 still had high survival rates. Most of the isolates and control isolates have strong tolerance to bile salts. The evaluation of fermentation characteristics indicated that the ability of the intestinal Lactobacillus to ferment skimmed milk was lower than that of the reference L. plantarum P8. In the process of storage, the viable count of screened isolates of human origin in fermented milk decreased to some extent, but remained above 7.01 ± 0.22 log CFU/ml, showing good storage characteristics.

Biofilm production ability and associated characteristics of Streptococcus agalactiae isolates from companion animals and humans

ObjectiveWe evaluated biofilm production ability (BPA) of Streptococcus agalactiae isolates from companion animals/humans and clarified the relationship between BPA populations and other microbiological features. MethodsCompanion animal-/human-origin isolates were collected with host information. We measured BPA using crystal violet staining, via virulence-associated gene profiling (hylB-pavA-pilB-spb1-srtC1-brpA), capsular genotyping, multilocus sequence typing, and antimicrobial resistance (AMR) phenotyping/genotyping. Significant difference in BPA of isolates from different hosts was assessed. We analyzed the association between BPA populations and the virulence genotypes, capsular genotypes, sequence types/clonal complexes, and AMR phenotypes/genotypes. Inhibitory effect of berberine on BPA was evaluated. ResultsFive, twenty-six, and twenty-six isolates belonged to strong, moderate, and weak biofilm producers, whereas seventeen showed no biofilm production. We defined strong, moderate, or weak biofilm producers as the producer group (n = 57) to conduct a comparative analysis between the producer and non-producer populations. There was a significant correlation between the producer population and vaginal specimen. We found significant associations between the producer group and presence (57.9%) of pilB and between the non-producer population and presence (70.6%) of spb1. There was no association between the producer group and capsular genotypes, sequence types/clonal complexes, and AMR phenotypes/genotypes (except for a significant correlation between the producer group and AMR to minocycline). We confirmed inhibitory effect of berberine at sub-minimum inhibitory concentrations (MICs) against the type strain on BPA. ConclusionOur observations suggest that S. agalactiae harboring pilB is more capable of producing biofilms, with berberine inhibitory effect at sub-MICs on BPA.

Genetic evaluation of Mycobacteriumbovis isolates with MIRU-VNTR and spoligotyping

Background/aimDetermining the epidemiological characteristics of M. bovis strains isolated from human and animal tuberculosis cases will assist in taking more appropriate and effective control measures in controlling tuberculosis originating from animals. Materials and methodsIn this study, 32 M. bovis isolates of animal origin and 10 of human origin were isolated and identified in the Çukurova region between March 2011 and June 2012. The 12-locus MIRU-VNTR and spoligotyping methods were used. ResultsSix different patterns were determined by spoligotyping and 10 by MIRU-VNTR. When both methods were used together, the number of patterns was found to be 28; MIRU4, MIRU26, MIRU31, and MIRU40 had the highest locus discrimination powers by MIRU-VNTR. The isolates concentrated in the SB0120 pattern at the rate of 42.85% in spoligotyping. By the same method, it was seen that 7 isolates were M bovis ssp. caprae pattern and 2 human isolates were M. bovis BCG pattern. Nevertheless, spoligotyping and MIRU-VNTR patterns showed that 5 M. bovis isolates of human origin were 100% compatible with isolates originating from cattle. ConclusionIn this study, we determined that the use of spoligotyping and MIRU-VNTR methods together was found to be more sensitive in the epidemiological analysis of M. bovis isolates.

Cross-sectional study of Staphyloccus lugdunensis prevalence in cats

Staphylococcus lugdunensis is a commensal bacterium in humans and other animals that can cause serious infections. The aim of this research was to estimate the frequency of S. lugdunensis in pet cats and to characterize the S. lugdunensis isolates obtained. The prevalence of S. lugdunensis was 0.77% (4/523) in healthy cats and 1.23% (1/81) in sick cats. The isolates (N = 5), which colonized conjunctival sacs, nares, and the anus, were almost fully phenotypically sensitive to antibiotics, but harbored resistance genes to four chemotherapeutic groups. Their sequence types (STs) included ST2, ST3, ST9, and ST15. There was detected a far lower prevalence of S. lugdunensis in pet cats than is reported in the human population. Nevertheless, the phenotypic and genotypic properties of S. lugdunensis isolates found in the current study were very similar to those described previously in isolates of human origin.

Molecular characteristics of oxazolidinone resistance in enterococci from a multicenter study in China

BackgroundLinezolid-resistant enterococci pose great challenges in clinical practice. The aim of this study is to study the mechanisms underlying the resistance and genetic environment of antimicrobial resistance gene of linezolid-resistant enterococci.ResultsThe linezolid MICs of 16 enterococci were 4 mg/L to 16 mg/L. Four strains belonged to multi-drug resistant (MDR) bacteria. The sequence types (STs) of 13 enterococci strains performed WGS were diverse: 3 ST476, 1 ST86, ST116, ST480, ST59, ST416, ST21, ST67, ST16, ST585 and ST18. None of them carried multi-drug resistance gene cfr. Only one strain had the G2658 T mutation of target 23S rRNA gene. Thirteen (13/16, 81.3%) strains harbored the novel oxazolidinone resistance gene optrA. WGS analysis showed that the optrA gene was flanked by sequence IS1216E insertion in 13 strains, and optrA was adjacent to transposons Tn558 in two strains and Tn554 in one strain. The optrA gene was identified to be co-localized with fexA, the resistance genes mediated florfenicol resistance in 13 strains, and ermA1, the resistance genes mediated erythromycin resistance in 9 strains, indicating that linezolid-resistant strains may be selected due to non-oxazolidinone antibiotics (i.e. macrolides and florfenicol) usage.ConclusionOur findings demonstrate the high diversity of optrA-carrying genetic platforms. The mobile genetic elements (MGEs) may play an important role in the dissemination of optrA into the enterococci isolates of human origin. The genetic evidence of transferable feature and co-selection of optrA should be gave more attention in clinical practice.

Presence of NDM in non-E. coli Enterobacteriaceae in the poultry production environment.

Characterization of non-Escherichia coli NDM-carrying Enterobacteriaceae in the poultry production environment. A total of 36 NDM-positive Enterobacteriaceae (22 Klebsiella pneumoniae, 13 Enterobacter cloacae and 1 Salmonella enterica) were isolated from a chicken farm and WGS was conducted using Illumina Hiseq2500. The genomic characterization of the isolates acquired through WGS analysis included the genomic context-flanking blaNDM genes, MLST, the antibiotic resistance genes (ARGs) and replicon types of plasmids. WGS information for another 73 K. pneumoniae isolates from different sources was retrieved from GenBank and then combined with isolates in this study for comparative genomic and phylogenetic analysis. Three types of genetic environment carrying blaNDM were identified in 36 non-E. coli Enterobacteriaceae isolates. Sequence comparison analysis indicated these genetic environments were completely identical to our previous findings. WGS further revealed three major types of plasmids (IncFIB, IncX3 and IncFII) from these isolates and the phylogenetic analysis suggested several K. pneumoniae isolates with ST11, ST37 and ST147 from the commercial chicken farm that were closely related to isolates of human origin. The blaNDM-harbouring genetic contexts were identified not only in E. coli, but also in K. pneumoniae, E. cloacae and S. enterica, which may indicate that blaNDM has been widely disseminated to non-E. coli Enterobacteriaceae species in animal farms. The close relationship of K. pneumoniae isolates from different origins suggests they could serve as a key vehicle for the transfer of ARGs between humans and food animal production environments.

Association of biofilm formation and cytotoxic potential with multidrug resistance in clinical isolates of Pseudomonas aeruginosa.

Multidrug resistant (MDR) Pseudomonas aeruginosa having strong biofilm potential and virulence factors are a serious threat for hospitalized patients having compromised immunity. In this study, 34 P. aeruginosa isolates of human origin (17 MDR and 17 non-MDR clinical isolates) were checked for biofilm formation potential in enriched and minimal media. The biofilms were detected using crystal violet method and a modified software package of the automated VideoScan screening method. Cytotoxic potential of the isolates was also investigated on HepG2, LoVo and T24 cell lines using automated VideoScan technology. Pulse field gel electrophoresis revealed 10 PFGE types in MDR and 8 in non-MDR isolates. Although all isolates showed biofilm formation potential, strong biofilm formation was found more in enriched media than in minimal media. Eight MDR isolates showed strong biofilm potential in both enriched and minimal media by both detection methods. Strong direct correlation between crystal violet and VideoScan methods was observed in identifying strong biofilm forming isolates. High cytotoxic effect was observed by 4 isolates in all cell lines used while 6 other isolates showed high cytotoxic effect on T24 cell line only. Strong association of multidrug resistance was found with biofilm formation as strong biofilms were observed significantly higher in MDR isolates (p-value < 0.05) than non-MDR isolates. No significant association of cytotoxic potential with multidrug resistance or biofilm formation was found (p-value > 0.05). The MDR isolates showing significant cytotoxic effects and strong biofilm formation impose a serious threat for hospitalized patients with weak immune system.

Screening and selection of probiotic Lactobacillus strains of Indian gut origin based on assessment of desired probiotic attributes combined with principal component and heatmap analysis

Selection of candidate probiotic strains for functional efficacy studies in cell line and animal models is an important challenge which requires a platform of basic information about their phenotype and genotype identity in relation to their probiotic attributes. Hence, in the current study, the probiotic isolates of human origin previously identified using PCR or 16S rRNA gene sequencing were characterized for probiotic phenotypes using FAO/WHO or ‘ICMR-DBT′ guidelines for evaluation of probiotics in food. Finally, the candidate probiotic strains were selected by subjecting their probiotic phenotypic data (acid tolerance, bile tolerance, cell surface hydrophobicity, aggregation, co-aggregation and antimicrobial activity) to multivariate principal component analysis and heatmap visualization. Out of the fourteen isolates, which included two L. casei (LbS2 and LbS6), four L. plantarum (A1, A3, A5 and A9) and eight L. fermentum (A2, A6, A7, A8, A10, LbS4, Ad05 and Ad06), a total of nine probiotic Lactobacillus isolates viz. A1, A2, A5, A6, A8, A9, Ad06, LbS2, LbS4 were finally selected as candidate probiotic strains.

Comparative analysis of an mcr-4 Salmonella enterica subsp. enterica monophasic variant of human and animal origin.

In this study we compared the recently described mcr-4-positive Salmonella enterica monophasic variant, isolated in 2016 in two Italian patients affected by gastroenteritis, with the first mcr-4-positive Salmonella isolate identified in 2013 in a pig at slaughter in Italy. WGS of the two Salmonella isolates of human origin was performed using a MiSeq instrument (Illumina). The phylogenetic analysis was performed by SNP analysis, comparing genomes of the mcr-4-positive isolates of swine and human origin with 82 Salmonella genomes downloaded from the EnteroBase Salmonella database. Complete sequences of plasmids carrying mcr-4.2 were obtained and compared. Transformation experiments were performed to transfer the mcr-4 plasmids into a colistin-susceptible Escherichia coli recipient strain. Comparative genomics demonstrated that the Salmonella of swine origin did not cluster with the isolates of human origin. The mcr-4.2 gene variant identified in the Salmonella of human origin was located on a ColE-like plasmid. This plasmid showed different replication and mobilization genes with respect to those previously described in the ColE plasmid carrying the mcr-4.1 variant, identified in Salmonella of swine origin. The divergence in genomes, plasmids and gene variants demonstrated that there was not a unique mcr-4-positive, monophasic Salmonella lineage circulating in animals and causing gastroenteritis in humans in Italy. There was no horizontal transfer of the same plasmid among Salmonella strains of animal and human origin, but the mcr-4 gene and a fragment of the plasmid identified in the animal strain were mobilized by an IS1294 into a different ColE plasmid.

In Vitro Activity of 3 Commercial Bacteriophage Cocktails Against Salmonella and Shigella spp. Isolates of Human Origin.

Background:Salmonella and Shigella spp. are 2 of the most frequent and deadly enteric bacterial pathogens recorded worldwide. In developing countries Salmonella infections are responsible for many deaths annually and these mortality rates are prone to increase due to the emergence of resistance to antibiotics. In this overall scenario new alternative therapeutic approaches are needed.Methods:For the first time, we investigated the activity of 3 commercial bacteriophage cocktails (INTESTI, Septaphage, PYO) against a collection of contemporary Salmonella spp. (n = 30) and Shigella spp. (n = 20) strains isolated in Switzerland. Phage susceptibility was determined by implementing the spot test.Results:The overall susceptibility of Salmonella spp. to INTESTI and Septaphage was 87% and 77%, respectively. With regard to Shigella spp., the overall susceptibility to INTESTI and Septaphage was 95% and 55%, respectively. PYO was observed to be active against only 10% of Salmonella spp. but against 95% of Shigella spp.Conclusions:Our results seem promising, especially for the INTESTI biopreparation against Salmonella enterica infections. Nevertheless, such speculation should be supported by further in vivo studies to confirm efficacy and safety of the cocktails. We also emphasize the importance of large in vitro screening analyses aimed to assess the activity of such biopreparations against contemporary multidrug-resistant strains that are emerging worldwide.

Characterisation of the Resistance Patterns to Non Beta-Lactam Antimicrobials in Esbl-Producing Enterobacteriaceae Isolated from Dogs and Their Owners

Enterobacteriaceae producing extended-spectrum beta-lactamase (ESBL) enzymes are resistant to beta-lactam agents and are also commonly multidrug resistant being associated with the resistance to other classes of antibiotics.The aim of our study was to characterise resistance patterns in non-beta-lactam antibiotics of ESBL-producing Enterobacteriaceae strains isolated from faecal matter of pets and owners.The study was carried out on 63 samples of faecal matter (42 from pets and 21 from owners). The ESBL screening was carried out using the Brilliance ESBL Oxoid chromogenic medium. The isolated strains that generated characteristic presumptive ESBL-producing colonies were cultivated on 5% sheep blood medium for the extraction of bacterial DNA using the boiled preps technique. The confirmation of E. coli species was performed molecularly based on the detection of blauidA and blauspA genes. Other Enterobacteriaceae species were identified based on the minimum biochemical characteristics using the MIU and TSI medium. The phenotypical confirmation of presumptive ESBL-producing strains was carried out using the Double Disc Synergy Test (DDST) using a combination of 3rd generation cephalosporins and beta-lactamase inhibitor agents. The determination of the resistance degree in other classes of antibiotics was carried out through the Kirby-Bauer diffusimetric method, and the results were interpreted according to the CLSI standard.Following the species investigation of isolates, 60/63 (95.28%) belonged to the E. coli species and 3/63 (4.72%) to the K. pneumoniae species. Animal isolates were resistant to sulphonamides (54.76% resistance to SXT), fluoroquinolones (45.23% resistance to ENR) and tetracyclines (54.75% resistance to TE). In addition to strains of animal origin for isolates of human origin, an increased resistance has been noticed to phenicols and aminoglycosides.This study has identified a high prevalence of ESBL-producing Enterobacteriaceae strains and associated with multidrug resistance for pets and their owners.

Detection of mcr-4 positive Salmonella enterica serovar Typhimurium in clinical isolates of human origin, Italy, October to November 2016.

In this study we report the detection of the recently described mcr-4 gene in two human isolates of Salmonella enterica serovar Typhimurium. The strains were isolated from faecal samples of two Italian patients with gastroenteritis, collected in 2016. The identified mcr-4 genes (variant mcr-4.2) differed from the mcr-4 gene originally described in a Salmonella strain of swine origin from Italy. Salmonella species could represent a hidden reservoir for mcr genes.

Antimicrobial Resistance in Nontyphoidal Salmonella Isolated from Human and Poultry-Related Samples in Brazil: 20-Year Meta-Analysis.

Nontyphoidal Salmonella are one of the leading causes of foodborne diseases in the world. As poultry products are recognized as main sources of human salmonellosis, nontyphoidal Salmonella control has become a global issue for the poultry industry. The increasing antimicrobial resistance in poultry-related nontyphoidal Salmonella serovars is a global matter of concern. By monitoring the evolution of antimicrobial resistance, alternative treatments can be identified and possible restrictions in the treatment of systemic human salmonellosis foreseen. A meta-analysis was conducted to assess the profile and temporal evolution of the antimicrobial resistance of nontyphoidal Salmonella of poultry and human origin in Brazil, isolated in the period from 1995 to 2014. Four databases were researched; twenty-nine articles met the eligibility criteria and were included in the meta-analysis. In the nontyphoidal isolates of poultry origin, the highest levels of antimicrobial resistance were verified for sulfonamides (44.3%), nalidixic acid (42.5%), and tetracycline (35.5%). In the human-origin isolates, the resistance occurred mainly for sulfonamides (46.4%), tetracycline (36.9%), and ampicillin (23.6%). Twenty-two articles described results of antimicrobial resistance specifically for Salmonella Enteritidis, also enabling the individual meta-analysis of this serovar. For most antimicrobials, the resistance levels of Salmonella Enteritidis were lower than those found when considering all the nontyphoidal serovars. In the poultry-origin isolates, a quadratic temporal distribution was observed, with reduced resistance to streptomycin in Salmonella Enteritidis and in all nontyphoidal serovars, and a linear increase of resistance to nalidixic acid in Salmonella Enteritidis. In the human-origin isolates, a linear increase was identified in the resistance to nalidixic acid in Salmonella Enteritidis and in all the nontyphoidal isolates, and to gentamicin in Salmonella Enteritidis. Continuous monitoring of the development and spread of antimicrobial resistance could support the measurement of the consequences on poultry and human health.

Multilocus sequence typing of Dientamoeba fragilis identified a major clone with widespread geographical distribution

The flagellated protozoan Dientamoeba fragilis is often detected in humans with gastrointestinal symptoms, but it is also commonly found in healthy subjects. As for other intestinal protozoa, the hypothesis that genetically dissimilar parasite isolates differ in their ability to cause symptoms has also been raised for D. fragilis. To date, only two D. fragilis genotypes (1 and 2) have been described, of which genotype 1 largely predominates worldwide. However, very few markers are available for genotyping studies and therefore the extent of genetic variation among isolates remains largely unknown. Here, we performed metagenomics experiments on two D. fragilis-positive stool samples, and identified a number of candidate markers based on sequence similarity to the phylogenetically related species Trichomonas vaginalis. Markers corresponding to structural genes and to genes encoding for proteases were selected for this study, and PCR experiments confirmed their belonging to the D. fragilis genome; two previously described markers (small subunit ribosomal DNA and large subunit of RNA polymerase II) were also included. Using this panel of markers, 111 isolates of human origin were genotyped, all of which, except one, belonged to genotype 1. These isolates had been collected at different times from symptomatic and asymptomatic persons of different age groups in Italy, Denmark, Brazil and Australia. By sequencing approximately 160kb from 500 PCR products, a very low level of polymorphism was observed across all the investigated loci, suggesting the existence of a major clone of D. fragilis with a widespread geographical distribution.

Evidence of Coxiella burnetii in Punjab province, Pakistan

Coxiella burnetii causes query (Q) fever, an important zoonotic disease with worldwide significance. The role of environment in the ecology of C. burnetti, and its influence on seroconversion in animals has not been elucidated in Pakistan. We carried out a cross-sectional study in Punjab province to (1) determine the prevalence and distribution of C. burnetii in soil using an ISIIII gene-based real time-polymerase chain reaction (RT-PCR) assay, (2) analyze association between the occurrence of C. burnetii in soil and its predictors i.e. soil characteristics (macro- and micro-nutrients) and several likely risk factors including the seroconversion in small ruminants at places where its genome had or had not been detected, and (3) predict homology and genetic diversity of the identified strains using sequences originated from different hosts worldwide. A total of 2425 soil samples from nine districts of Punjab province were processed. C. burnetii DNA was detected in 47 samples (1.94%, 95% CI: ±0.55) originating from 35 villages of studied districts (7.22%, 95% CI: ±2.30). The highest prevalence was found in Attock (7.11%, 95% CI: ±3.36), followed by Lahore (4.83%, 95% CI: ±3.49), Sahiwal (4.70%, 95% CI: ±2.6), Dera Ghazi Khan (2.33%, 95% CI: ±2.02), Faisalabad (1.35%, 95% CI: ±1.18) and Sheikhupura (0.68%, 95% CI: ±0.94). The odds of detecting bacterial DNA in soil was increased with a unit increase in organic matter [2.511 (95% CI: 1.453–4.340), p=0.001] and sodium [1.013 (95% CI: 1.005–1.022), p=0.001], whereas, calcium [0.984 (95% CI: 0.975–0.994), p=0.002] and potassium [0.994 (95% CI: 0.990–0.999), p=0.011] had protective effect where a unit increase in each analyte decreased odds for its occurrence by 1.0% approximately. Likewise, for categorical variables (risk factors), the odds of detecting C. burnetii were higher at locations >500m away from a main road [1.95 (95% CI: 1.06–3.78), p=0.04]. The enzyme-linked immunosorbent assay (ELISA) revealed an increased prevalence of antibodies in sheep (17.9%, 95% CI: ±5.54) compared with goats (16.4%, 95% CI: ±4.34). When determining the association between soil DNA and C. burnetii antibodies in small ruminants, the odds of detecting these antibodies were significant in sheep at the livestock barns [2.81 (95% CI: 1.20–7.37), p=0.02]. The IS1111 gene-based sequence analysis revealed a clustering of the DNA into two distinct groups with much genetic divergence (0.76–68.70%): the first group that contained sequences from Lahore district clustered with human and buffalo origin isolates, whereas the second group that contained the sequences from the remaining study districts clustered with goat-, rodent- and human-origin isolates. This study provides the first evidence of the presence of C. burnetii in the environment in Punjab province, Pakistan. Future studies are needed to ascertain the bacteria’s molecular epidemiology over a wide geographical area, type the isolates, and evaluates the potential risks to human populations, particularly farmers and veterinarians.