Nonhuman Strains Research Articles

Characteristics of Clinical Isolates of Listeria monocytogenes in Sichuan, China, in 2022 Based on Whole Genome Sequencing Analysis.

Listeria monocytogenes is a foodborne pathogen. In 2022, we collected 15 strains of L. monocytogenes isolated from patients in some foodborne disease sentinel monitoring hospitals in Sichuan Province. Through whole genome sequencing (WGS), we obtained the virulence genes carried by the strains, multi-locus sequence typing (MLST), core genome MLST (cgMLST), clonal complex (CC), and serum groups and constructed a phylogenetic tree and minimum spanning tree with nonhuman strains. An analysis shows that all 15 strains of L. monocytogenes carry virulence genes LIPI-1 and LIPI-2, whereas the carrying rates of LIPI-3 and LIPI-4 virulence genes are relatively low. The MLST typing results showed a total of 10 sequence types (ST), including 10 CCs, with ST7 being the dominant type. The cgMLST clearly distinguishes strains of different lineages and CC types. The serum group is divided into three types: IIa, IIb, and IVb, with IIa being the dominant serum group. An analysis of antibiotic genes showed that all 15 strains carried FosX, lin, mprF, and norB with high carrying rates. The minimum inhibitory concentration results indicated that all were susceptible to eight antibiotics (ampicillin, penicillin, tetracycline, meropenem, erythromycin, vancomycin, ciprofloxacin, and trimethoprim-sulfamethoxazole). The analysis of strains isolated from different sources of Listeria revealed varying degrees of diversity, and the contamination of meat and environment within the province is closely related to clinical cases. L. monocytogenes isolated from clinical cases in Sichuan Province carry multiple virulence and antibiotic genes, with high potential pathogenicity. It is necessary to further strengthen the monitoring and control of food and environment by L. monocytogenes within Sichuan Province.

Phylogenomic and phenotypic analyses highlight the diversity of antibiotic resistance and virulence in both human and non-human Acinetobacter baumannii.

Acinetobacter baumannii is a Gram-negative, opportunistic pathogen that causes infections in the immunocompromised. With a high incidence of muti-drug resistance, carbapenem-resistant A. baumannii is designated as a priority 1 pathogen by the WHO. The current literature has expertly characterized clinical isolates of A. baumannii. As the challenge of these infections has recently been classified as a One Health issue, we set out to explore the diversity of isolates from human and non-clinical sources, such as agricultural surface water, urban streams, various effluents from wastewater treatment plants, and food (tank milk); and, importantly, these isolates came from a wide geographic distribution. Phylogenomic analysis considering almost 200 isolates showed that our diverse set is well-differentiated from the main international clones of A. baumannii. We discovered novel sequence types in both hospital and non-clinical settings and five strains that overexpress the resistance-nodulation-division efflux pump adeIJK without changes in susceptibility reflected by this overexpression. Furthermore, we detected a bla ADC-79 in a non-human isolate despite its sensitivity to all antibiotics. There was no significant differentiation between the virulence profiles of clinical and non-clinical isolates in the Galleria mellonella insect model of virulence, suggesting that virulence is neither dependent on geographic origin nor isolation source. The detection of antibiotic resistance and virulence genes in non-human strains suggests that these isolates may act as a genetic reservoir for clinical strains. This endorses the notion that in order to combat multi-drug-resistant infection caused by A. baumannii, a One Health approach is required, and a deeper understanding of non-clinical strains must be achieved.IMPORTANCEThe global crisis of antibiotic resistance is a silent one. More and more bacteria are becoming resistant to all antibiotics available for treatment, leaving no options remaining. This includes Acinetobacter baumannii. This Gram-negative, opportunistic pathogen shows a high frequency of multi-drug resistance, and many strains are resistant to the last-resort drugs carbapenem and colistin. Research has focused on strains of clinical origin, but there is a knowledge gap regarding virulence traits, particularly how A. baumannii became the notorious pathogen of today. Antibiotic resistance and virulence genes have been detected in strains from animals and environmental locations such as grass and soil. As such, A. baumannii is a One Health concern, which includes the health of humans, animals, and the environment. Thus, in order to truly combat the antibiotic resistance crisis, we need to understand the antibiotic resistance and virulence gene reservoirs of this pathogen under the One Health continuum.

Antimicrobial Resistance and Molecular Characterization of Salmonella Rissen Isolated in China During 2008-2019.

This study aimed to provide epidemiological features of Salmonella enterica serovar Rissen, determine antimicrobial susceptibility, virulence gene profiles, and describe the potential association of S. Rissen from different sources in China. During 2008-2019, a total of non-repetitive 228 S. Rissen isolates were collected from human, animals and environment in China. The antimicrobial susceptibility test, screening of antimicrobial and virulence genes by PCR, and pulsed-field gel electrophoresis (PFGE) were performed. Among the 154 isolates from human, the majority of the cases (80.5%) occurred in summer, and S. Rissen was mainly detected in people aged 21-40 (37.7%) and 41-60 (28.6%) years old, and 74 non-human source S. Rissen strains were identified, with pork being the most common source. About 93.4% isolates were resistant to at least one of the 12 tested antimicrobial agents, and high frequencies of resistance were observed for tetracyclines (91.2%), trimethoprim-sulfamethoxazole (74.1%) and ampicillin (67.5%). A total of 171 (75%) isolates were resistant to at least three categories of antimicrobials, and the most common resistance profile was Tetracycline(s)-β-Lactams-Sulfonamides. The resistance rates to chloramphenicol, quinolones and sulfafurazole were significantly higher in strains isolated from human compared to non-human source strains. Among these isolates, the β-Lactams resistance was mainly associated with gene blaTEM (54.7%), sulfonamide resistance with sul2 (45.7%) and sul3 (54.3%), tetracycline resistance with tetA (81.3%). All the isolates harbored virulence genes hilA, sopB, sciN, stn and ssrB, and most of them harbored ssaQ (98.7%), mgtC (98.7%) and invA (98.2%). The majority (91.7%) of S. Rissen isolates showed high similarity (>80%) with each other in PFGE patterns and came from human, animals and environment. The high frequencies of multidrug resistance and probable clonal dissemination in this serovar call for the necessity of systematic surveillance on S. Rissen in China.

Human left ventricular cardiac fibroblasts undergo a dynamic shift in secretome and gene expression toward a cardiac myofibroblast phenotype during early passage in typical culture expansion conditions

Cardiac fibroblasts (CFs) are critical components of the cardiac niche and primarily responsible for assembly and maintenance of the cardiac extracellular matrix (ECM). CFs are increasingly of interest for tissue engineering and drug development applications, as they provide synergistic support to cardiomyocytes through direct cell-to-cell signaling and cell-to-ECM interactions via soluble factors, including cytokines, growth factors and extracellular vesicles. CFs can be activated to a cardiac myofibroblast (CMF) phenotype upon injury or stimulation with transforming growth factor beta 1. Once activated, CMFs assemble collagen-rich ECM, which is vitally important to stabilize scar formation following myocardial infarction, for example. Although there is greater experience with culture expansion of CFs among non-human strains, very little is known about human CF-to-CMF transitions and expression patterns during culture expansion. In this study, we evaluated for shifts in inflammatory and angiogenic expression profiles of human CFs in typical culture expansion conditions. Understanding shifts in cellular expression patterns during CF culture expansion is critically important to establish quality benchmarks and optimize large-scale manufacturing for future clinical applications.

Detecting zoonotic Influenza A using QIAstat-Dx Respiratory SARS-CoV-2 panel for pandemic preparedness

Recent reports from the World Health Organization regarding Influenza A cases of zoonotic origin in humans (H1v and H9N2) and publications describing emergence swine Influenza A cases in humans together with “G4” Eurasian avian-like H1N1 Influenza A virus have drawn global attention to Influenza A pandemic threat. Additionally, the current COVID-19 epidemic has stressed the importance of surveillance and preparedness to prevent potential outbreaks. One feature of the QIAstat-Dx Respiratory SARS-CoV-2 panel is the double target approach for Influenza A detection of seasonal strains affecting humans using a generic Influenza A assay plus the three specific human subtype assays. This work explores the potential use of this double target approach in the QIAstat-Dx Respiratory SARS-Co-V-2 Panel as a tool to detect zoonotic Influenza A strains. A set of recently recorded H9 and H1 spillover strains and the G4 EA Influenza A strains as example of recent zoonotic Flu A strains were subjected to detection prediction with QIAstat-Dx Respiratory SARS-CoV-2 Panel using commercial synthetic dsDNA sequences. In addition, a large set of available commercial human and non-human influenza A strains were also tested using QIAstat-Dx Respiratory SARS-CoV-2 Panel for a better understanding of detection and discrimination of Influenza A strains. Results show that QIAstat-Dx Respiratory SARS-CoV-2 Panel generic Influenza A assay detects all the recently recorded H9, H5 and H1 zoonotic spillover strains and all the G4 EA Influenza A strains. Additionally, these strains yielded negative results for the three-human seasonal IAV (H1, H3 and H1N1 pandemic) assays. Additional non-human strains corroborated those results of Flu A detection with no subtype discrimination, whereas human Influenza strains were positively discriminated. These results indicate that QIAstat-Dx Respiratory SARS-CoV-2 Panel could be a useful tool to diagnose zoonotic Influenza A strains and differentiate them from the seasonal strains commonly affecting humans.

Comparison and selection of probiotic Lactobacillus from human intestinal tract and traditional fermented food in vitro via PCA, unsupervised clustering algorithm, and heat-map analysis.

Traditional fermented products and human intestines are rich sources of Lactobacillus strains which may have remarkable probiotic properties. In the present study, the probiotic properties of 40 Lactobacillus strains isolated from intestinal tracts of longevity population and traditional fermented food in China were determined, including the survival rates in simulated gastric acid and bile salt, aggregation, hydrophobicity, adhesion rate, antioxidant ability (ferric reducing antioxidant power), and antimicrobial ability. The differences between human strains and nonhuman strains were compared via t-test and principal component analysis (PCA). The significant differences were found in the survival rate at 0.3% bile salt, adhesion ability of the strains, and antioxidant ability of the fermentation broth (p < .05). The results of PCA showed that the first principal component (PC1) score of human strains was significantly higher than that of nonhuman strains (p < .01). And some probiotic Lactobacillus were selected for further application based on the unsupervised clustering algorithm, heat-map analysis, and K-means algorithm. Four strains, CS128, CS39, CS01, and CS1301, along with Lactobacillus rhamnosus GG (LGG) were divided into cluster I. The four strains, all isolated from human tracts, have been selected. Thus, human Lactobacillus has better probiotic potential and application prospects than strains from the nonhuman source. PCA, the unsupervised clustering algorithm, and heat-map analysis can be used to analyze and select Lactobacillus visually and effectively.

Signatures of hybridization in Trypanosoma brucei.

Genetic exchange among disease-causing micro-organisms can generate progeny that combine different pathogenic traits. Though sexual reproduction has been described in trypanosomes, its impact on the epidemiology of Human African Trypanosomiasis (HAT) remains controversial. However, human infective and non-human infective strains of Trypanosoma brucei circulate in the same transmission cycles in HAT endemic areas in subsaharan Africa, providing the opportunity for mating during the developmental cycle in the tsetse fly vector. Here we investigated inheritance among progeny from a laboratory cross of T. brucei and then applied these insights to genomic analysis of field-collected isolates to identify signatures of past genetic exchange. Genomes of two parental and four hybrid progeny clones with a range of DNA contents were assembled and analysed by k-mer and single nucleotide polymorphism (SNP) frequencies to determine heterozygosity and chromosomal inheritance. Variant surface glycoprotein (VSG) genes and kinetoplast (mitochondrial) DNA maxi- and minicircles were extracted from each genome to examine how each of these components was inherited in the hybrid progeny. The same bioinformatic approaches were applied to an additional 37 genomes representing the diversity of T. brucei in subsaharan Africa and T. evansi. SNP analysis provided evidence of crossover events affecting all 11 pairs of megabase chromosomes and demonstrated that polyploid hybrids were formed post-meiotically and not by fusion of the parental diploid cells. VSGs and kinetoplast DNA minicircles were inherited biparentally, with approximately equal numbers from each parent, whereas maxicircles were inherited uniparentally. Extrapolation of these findings to field isolates allowed us to distinguish clonal descent from hybridization by comparing maxicircle genotype to VSG and minicircle repertoires. Discordance between maxicircle genotype and VSG and minicircle repertoires indicated inter-lineage hybridization. Significantly, some of the hybridization events we identified involved human infective and non-human infective trypanosomes circulating in the same geographic areas.

The epidemiology of Plasmodium vivax among adults in the Democratic Republic of the Congo

Reports of P. vivax infections among Duffy-negative hosts have accumulated throughout sub-Saharan Africa. Despite this growing body of evidence, no nationally representative epidemiological surveys of P. vivax in sub-Saharan Africa have been performed. To overcome this gap in knowledge, we screened over 17,000 adults in the Democratic Republic of the Congo (DRC) for P. vivax using samples from the 2013-2014 Demographic Health Survey. Overall, we found a 2.97% (95% CI: 2.28%, 3.65%) prevalence of P. vivax infections across the DRC. Infections were associated with few risk-factors and demonstrated a relatively flat distribution of prevalence across space with focal regions of relatively higher prevalence in the north and northeast. Mitochondrial genomes suggested that DRC P. vivax were distinct from circulating non-human ape strains and an ancestral European P. vivax strain, and instead may be part of a separate contemporary clade. Our findings suggest P. vivax is diffusely spread across the DRC at a low prevalence, which may be associated with long-term carriage of low parasitemia, frequent relapses, or a general pool of infections with limited forward propagation.

Patterns of resistance to antimicrobials in serovars of Salmonella enterica in Peru, 2012-2015

Salmonellosis is a universal zoonosis, causing frequent outbreaks of foodborne illness; Salmonella enterica is the species with the highest prevalence, a progressive increase in its resistance to antimicrobials is described. To determine the frequency of serovars and antimicrobial resistance patterns in S. enterica isolates submitted to the National Institute of Health, Lima, Peru. This is a cross-sectional study. All strains referred as part of national laboratory-based surveillance between 2012 and 2015 were included in the study. Strains were confirmed by conventional tests and serotyped by the Kauffmann-White scheme; antimicrobial susceptibility and confirmation of the BLEE phenotype was performed according to the method of Kirby-Bauer and Jarlier's method. A total of 540 strains of S. enterica were included in the study, where 96% (520/540) corresponded to human strains and 4% (20/540) to non-human strains (birds, food and environmental). In human samples, the most frequent serovar was S. Infantis (57%), followed by S. Enteritidis (27%) and S. Typhimurium (6%). High resistance to nitrofurantoin (74%), nalidixic acid (64%), ciprofloxacin (63%), tetracycline (63%), ampicillin (56%), sulfamethoxazole-trimethoprim (56%), cefotaxime (53%) and chloramphenicol (50%) was detected. In non-human samples, the most frequent serotype was S. Infantis (45%), followed by S. Typhimurium (40%) and S. Enteritidis (10%); a high resistance to nalidixic acid (55%), ciprofloxacin (45%), sulfamethoxazole-trimethoprim (40%), nitrofurantoin (40%), tetracycline (40%) was found. 65% of all strains had resistance to more than two antibiotics, 43,3% were ESBL producers and 99% of these had resistance between six and eight antibiotics. We found a high frequency of S. Infantis producing ESBL with multi-resistance to the antimicrobials in human and nonhuman samples received by the National Institute of Health.

Retrospective use of whole genome sequencing to better understand an outbreak of Salmonella enterica serovar Mbandaka in New South Wales, Australia.

IntroductionSalmonella enterica serovar Mbandaka is an infrequent cause of salmonellosis in New South Wales (NSW) with an average of 17 cases reported annually. This study examined the added value of whole genome sequencing (WGS) for investigating a non-point source outbreak of Salmonella ser. Mbandaka with limited geographical spread.MethodsIn February 2016, an increase in Salmonella ser. Mbandaka was noted in New South Wales, and an investigation was initiated. A WGS study was conducted three months after the initial investigation, analysing the outbreak Salmonella ser. Mbandaka isolates along with 17 human and non-human reference strains from 2010 to 2015.ResultsWGS analysis distinguished the original outbreak cases (n = 29) into two main clusters: Cluster A (n = 11) and Cluster B (n = 6); there were also 12 sporadic cases. Reanalysis of food consumption histories of cases by WGS cluster provided additional specificity when assessing associations.DiscussionWGS has been widely acknowledged as a promising high-resolution typing tool for enteric pathogens. This study was one of the first to apply WGS to a geographically limited cluster of salmonellosis in Australia. WGS clearly distinguished the outbreak cases into distinct clusters, demonstrating its potential value for use in real time to support non-point source foodborne disease outbreaks of limited geographical spread.

Effect of microorganisms on reducing cadmium uptake and toxicity in rice (Oryza sativa L.).

This study analyzed the application of three microorganism inoculums, including Bacillus subtilis, Bacillus cereus, and commercial effective microorganism (EM) solution in order to determine cadmium (Cd) reduction in rice (Oryza sativa L.) and rice growth promotion. Rice was grown in Cd-contaminated soil (120mg/kg) and selected microorganisms were inoculated. Cd concentration and rice weight were measured at 45 and 120days of the experiment. The result showed that B. subtilis inoculation into rice can highly reduce Cd accumulation in every part of rice roots and shoots (45days), and grains (120days). This species can effectively absorb Cd compared to other inoculums, which might be the main mechanism to reduce Cd transportation in rice plants. Interestingly, plants that were inoculated with bacterial species individually harbored higher calcium (Ca) and magnesium (Mg) accumulation; B. subtilis-inoculated plants had the highest levels of Ca and Mg compared to other inoculated ones. Moreover, inoculating rice plants with these microorganisms could increase the dry weight of the plant and protect them from Cd stress because all the inoculums presented the ability to solubilize phosphate, produce indole-3-acetic acid (IAA), and control ethylene levels by 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity. After 120days, quantification of each inoculum by quantitative polymerase chain reaction (qPCR) confirmed the root colonization of bacterial species, with B. subtilis showing higher 16S rRNA gene copy numbers than the other species. B. subtilis was classified as a non-human pathogenic strain, reassuring the safe application of this plant growth-promoting bacterium as a crop inoculum.

Comparative Genomics of Herpesviridae Family to Look for Potential Signatures of Human Infecting Strains

Herpesviridae family is one of the significant viral families which comprises major pathogens of a wide range of hosts. This family includes at least eight species of viruses which are known to infect humans. This family has evolved 180–220 million years ago and the present study highlights that it is still evolving and more genes can be added to the repertoire of this family. In addition, its core-genome includes important viral proteins including glycoprotein B and helicase. Most of the infections caused by human herpesviruses have no definitive cure; thus, search for new therapeutic strategies is necessary. The present study finds core-genome of human herpesviruses that differs from that of Herpesviridae family and nonhuman herpes strains of this family and might be a putative target for vaccine development. The phylogenetic reconstruction based upon the protein sequences of core gene set of Herpesviridae family reveals the sharp splits of its different subfamilies and supports the hypothesis of coevolution of viruses with their hosts. In addition, data mining for cis-elements in the genomes of human herpesviruses results in the prediction of numerous regulatory elements which can be used for regulating the expression of viral based vectors implicated in gene therapies.

Collagen-like proteins (ClpA, ClpB, ClpC, and ClpD) are required for biofilm formation and adhesion to plant roots by Bacillus amyloliquefaciens FZB42.

The genes of collagen-like proteins (CLPs) have been identified in a broad range of bacteria, including some human pathogens. They are important for biofilm formation and bacterial adhesion to host cells in some human pathogenic bacteria, including several Bacillus spp. strains. Interestingly, some bacterial CLP-encoding genes (clps) have also been found in non-human pathogenic strains such as B. cereus and B. amyloliquefaciens, which are types of plant-growth promoting rhizobacteria (PGPR). In this study, we investigated a putative cluster of clps in B. amyloliquefaciens strain FZB42 and a collagen-related structural motif containing glycine-X-threonine repeats was found in the genes RBAM_007740, RBAM_007750, RBAM_007760, and RBAM_007770. Interestingly, biofilm formation was disrupted when these genes were inactivated separately. Scanning electron microscopy and hydrophobicity value detection were used to assess the bacterial cell shape morphology and cell surface architecture of clps mutant cells. The results showed that the CLPs appeared to have roles in bacterial autoaggregation, as well as adherence to the surface of abiotic materials and the roots of Arabidopsis thaliana. Thus, we suggest that the CLPs located in the outer layer of the bacterial cell (including the cell wall, outer membrane, flagella, or other associated structures) play important roles in biofilm formation and bacteria-plant interactions. This is the first study to analyze the function of a collagen-like motif-containing protein in a PGPR bacterium. Knocking out each clp gene produced distinctive morphological phenotypes, which demonstrated that each product may play specific roles in biofilm formation. Our in silico analysis suggested that these four tandemly ranked genes might not belong to an operon, but further studies are required at the molecular level to test this hypothesis. These results provide insights into the functions of clps during interactions between bacteria and plants.

Non-avian animal reservoirs present a source of influenza A PB1-F2 proteins with novel virulence-enhancing markers.

PB1-F2 protein, expressed from an alternative reading frame of most influenza A virus (IAV) PB1 segments, may possess specific residues associated with enhanced inflammation (L62, R75, R79, and L82) and cytotoxicity (I68, L69, and V70). These residues were shown to increase the pathogenicity of primary viral and secondary bacterial infections in a mouse model. In contrast to human seasonal influenza strains, virulence-associated residues are present in PB1-F2 proteins from pandemic H1N1 1918, H2N2 1957, and H3N2 1968, and highly pathogenic H5N1 strains, suggesting their contribution to viruses' pathogenic phenotypes. Non-human influenza strains may act as donors of virulent PB1-F2 proteins. Previously, avian influenza strains were identified as a potential source of inflammatory, but not cytotoxic, PB1-F2 residues. Here, we analyze the frequency of virulence-associated residues in PB1-F2 sequences from IAVs circulating in mammalian species in close contact with humans: pigs, horses, and dogs. All four inflammatory residues were found in PB1-F2 proteins from these viruses. Among cytotoxic residues, I68 was the most common and was especially prevalent in equine and canine IAVs. Historically, PB1-F2 from equine (about 75%) and canine (about 20%) IAVs were most likely to have combinations of the highest numbers of residues associated with inflammation and cytotoxicity, compared to about 7% of swine IAVs. Our analyses show that, in addition to birds, pigs, horses, and dogs are potentially important sources of pathogenic PB1-F2 variants. There is a need for surveillance of IAVs with genetic markers of virulence that may be emerging from these reservoirs in order to improve pandemic preparedness and response.

African monkeys are infected by Plasmodium falciparum nonhuman primate-specific strains

Recent molecular exploration of the Plasmodium species circulating in great apes in Africa has revealed the existence of a large and previously unknown diversity of Plasmodium. For instance, gorillas were found to be infected by parasites closely related to Plasmodium falciparum, suggesting that the human malignant malaria agent may have arisen after a transfer from gorillas. Although this scenario is likely in light of the data collected in great apes, it remained to be ascertained whether P. falciparum-related parasites may infect other nonhuman primates in Africa. Using molecular tools, we here explore the diversity of Plasmodium species infecting monkeys in Central Africa. In addition to previously described Hepatocystis and Plasmodium species (Plasmodium gonderi and Plasmodium sp DAJ-2004), we have found one African monkey to be infected by a P. falciparum-related parasite. Examination of the nuclear and mitochondrial genomes of this parasite reveals that it is specific of nonhuman primates, indicating that P. falciparum-related pathogens can naturally circulate in some monkey populations in Africa. We also show that at least two distinct genetic entities of P. falciparum infect nonhuman primates and humans, respectively. Our discoveries bring into question the proposed gorilla origin of human P. falciparum.

Detection of virulent Escherichia coli O157 strains using multiplex PCR and single base sequencing for SNP characterization

To compare 167 Norwegian human and nonhuman Escherichia coli O157:H7/NM (nonmotile) isolates with respect to an A/T single nucleotide polymorphism (SNP) in the tir gene and to detect specific SNPs that differentiate STEC O157 into distinct virulence clades (1-3 and 8). We developed a multiplex PCR followed by single base sequencing for detection of the SNPs, and examined the association among SNP genotype, virulence profile (stx and eae status), multilocus variable number of tandem repeats analysis (MLVA) profile and clinical outcome. We found an over-representation of the T allele among human strains compared to nonhuman strains, including 5/6 haemolytic-uraemic syndrome cases. Fourteen strains belonged to clade 8, followed by two clade 2 strains. No clade 1 nor 3 isolates were observed. stx1 in combination with either stx2(EDL933) or stx2c were frequently observed among human strains, whereas stx2c was dominating in nonhuman strains. MLVA indicated that only single cases or small outbreaks with E. coli O157 have been observed in Norway through the years 1993-2008. We observed that the tir-255 A/T SNP and the stx status were different between human and nonhuman O157 strains. No major outbreaks were observed, and only a few strains were differentiated into the virulence clades 2 and 8. The detection of virulence clade-specific SNPs enables the rapid designation of virulent E. coli O157 strains, especially in outbreak situations.

Animal and human pathogenic Escherichia coli strains share common genetic backgrounds

Escherichia coli is a versatile species encompassing both commensals of the digestive tracts of many vertebrates, including humans, and pathogenic strains causing various intra- and extraintestinal infections. Despite extensive gene flow between strains, the E. coli species has a globally clonal population structure, consisting of distinct phylogenetic groups. Little is known about the relationships between phylogenetic groups and host specificity. We therefore used multilocus sequence typing (MLST) to investigate phylogenetic relationships and evaluated the virulence gene content of 35 E. coli strains representative of the diverse diseases encountered in domestic animals. We compared these strains with a panel of 101 human pathogenic and 98 non-human and human commensal strains representative of the phylogenetic and pathovar diversity of this species. A global factorial analysis of correspondence indicated that extraintestinal infections were caused mostly by phylogenetic group B2 strains, whereas intraintestinal infections were caused mostly by phylogenetic group A/B1/E strains, with strains responsible from extraintestinal or intraintestinal infections having specific virulence factors. It was not possible to distinguish between strains of human and animal origin. A detailed phylogenetic analysis of the MLST data showed that numerous pathogenic animal and human strains are very closely related, and had a number of virulence genes in common. However, a set of specific adhesins was identified in animal non-B2 group strains of all pathotypes. In conclusion, human and animal pathogenic strains share common genetic backgrounds, but non-B2 strains of different origins seem to have different sets of adhesins that could be involved in host specificity.

Characterization of genotype G8 strains from Malawi, Kenya, and South Africa

Reviews of the global distribution of rotavirus genotypes have revealed the continuous circulation of G8 strains in Africa, often responsible for more cases of rotavirus disease than the more common G1-G4 rotavirus strains. During the study, genotype G8 strains from Malawi, Kenya, and South Africa were detected and the VP7 and VP4 genes of selected specimens were sequenced. Results indicated that G8 strains appeared to reassort frequently and were associated with P[6], P[4], and P[8] specificity. Phylogenetic analysis suggested that G8 strains occurred in a North/South African phylogenetic divide. In addition, G8 strains appear to be able to infect non-human primates and strains with close phylogenetic relationships were detected in the same year on two continents. Any rotavirus vaccine introduced into African environments will need to demonstrate protective efficacy against unusual genotype combinations, new serotypes, and animal strains. Therefore, continuous monitoring of rotavirus strains in human and animal populations in Africa is a necessity.

Major histocompatibility complex class I binding predictions as a tool in epitope discovery

Over the last decade, in silico models of the major histocompatibility complex (MHC) class I pathway have developed significantly. Before, peptide binding could only be reliably modelled for a few major human or mouse histocompatibility molecules; now, high-accuracy predictions are available for any human leucocyte antigen (HLA) -A or -B molecule with known protein sequence. Furthermore, peptide binding to MHC molecules from several non-human primates, mouse strains and other mammals can now be predicted. In this review, a number of different prediction methods are briefly explained, highlighting the most useful and historically important. Selected case stories, where these 'reverse immunology' systems have been used in actual epitope discovery, are briefly reviewed. We conclude that this new generation of epitope discovery systems has become a highly efficient tool for epitope discovery, and recommend that the less accurate prediction systems of the past be abandoned, as these are obsolete.

Investigation of Lymphocyte Depletion and Repopulation Using Alemtuzumab (Campath-1H) in Cynomolgus Monkeys

As the target CD52 molecule is expressed on erythrocytes of most nonhuman primate strains, using alemtuzumab in these species would cause massive hemolysis. Six cynomolgus monkeys of Indonesian origin, screened by agglutination assay for absence of CD52 on erythrocytes, were administered alemtuzumab in a cumulative dose to a maximum of 60 mg/kg. In two monkeys, mycophenolate mofetil (MMF) was added as maintenance therapy. Complete depletion of T and B lymphocytes (>99.5%) was achieved with 20 mg/kg alemtuzumab and was more profound than in monkeys treated with antithymocyte globulin (n = 5), as quantified by flow cytometry. Repopulation was suppressed by weekly injections of 10 mg/kg. Without MMF, repopulation of CD20(+)B cells and CD8(+)T cells was complete within 2 and 3 months, respectively, and repopulation of CD4(+)T cells was 67% after 1 year. MMF significantly delayed CD4(+)T-cell repopulation. Among repopulating CD4(+) and CD8(+) T cells, a phenotypic shift was observed from CD45RA(hi)CD62L(hi) naïve cells toward CD45RA(lo)CD62L(lo) effector memory cells. In lymph nodes, the depletion of naïve cells was more profound than of memory cells, which may have initiated a proliferation of memory cells. This model offers opportunities to investigate lymphocyte depletion/repopulation phenomena, as well as the efficacy of alemtuzumab in preclinical transplantation models.