Phylogenetic and Evolutionary Relationships among Yellow Fever Virus Isolates in Africa

Previous studies with a limited number of strains have indicated that there are two genotypes of yellow fever (YF) virus in Africa, one in west Africa and the other in east and central Africa. We have examined the prM/M and a portion of the E protein for a panel of 38 wild strains of YF virus from Africa representing different countries and times of isolation. Examination of the strains revealed a more complex genetic relationship than previously reported. Overall, nucleotide substitutions varied from 0 to 25.8% and amino acid substitutions varied from 0 to 9.1%. Phylogenetic analysis using parsimony and neighbor-joining algorithms identified five distinct genotypes: central/east Africa, east Africa, Angola, west Africa I, and west Africa II. Extensive variation within genotypes was observed. Members of west African genotype II and central/east African genotype differed by 2.8% or less, while west Africa genotype I varied up to 6.8% at the nucleotide level. We speculate that the former two genotypes exist in enzootic transmission cycles, while the latter is genetically more heterogeneous due to regular human epidemics. The nucleotide sequence of the Angola genotype diverged from the others by 15.7 to 23.0% but only 0.4 to 5.6% at the amino acid level, suggesting that this genotype most likely diverged from a progenitor YF virus in east/central Africa many years ago, prior to the separation of the other east/central African strains analyzed in this study, and has evolved independently. These data demonstrate that there are multiple genotypes of YF virus in Africa and suggest independent evolution of YF virus in different areas of Africa.     

Genetic relationships and evolution of genotypes of yellow fever virus and other members of the yellow fever virus group within the Flavivirus genus based on the 3' noncoding region

Genetic relationships among flaviviruses within the yellow fever (YF) virus genetic group were investigated by comparing nucleotide sequences of the 3' noncoding region (3'NCR). Size heterogeneity was observed between members and even among strains of the same viral species. Size variation between YF strains was due to duplications and/or deletions of repeated nucleotide sequence elements (RYF). West African genotypes had three copies of the RYF (RYF1, RYF2, and RYF3); the Angola and the East and Central African genotypes had two copies (RYF1 and RYF3); and South American genotypes had only a single copy (RYF3). Nucleotide sequence analyses suggest a deletion within the 3'NCR of South American genotypes, including RYF1 and RYF2. Based on studies with the French neurotropic vaccine strain, passage of a YF virus strain in cell culture can result in deletion of RYF1 and RYF2. Taken together, these observations suggest that South American genotypes of YF virus evolved from West African genotypes and that the South American genotypes lost RYF1 and RYF2, possibly in a single event. Repeated sequence elements were found within the 3'NCR of other members of the YF virus genetic group, suggesting that it is probably characteristic for members of the YF virus genetic group. A core sequence of 15 nucleotides, containing two stem-loops, was found within the 3'NCR of all members of the YF genetic group and may represent the progenitor repeat sequence. Secondary structure predictions of the 3'NCR showed very similar structures for viruses that were closely related phylogenetically.     

Genetic Diversity of Newcastle Disease Virus in Wild Birds and Pigeons in West Africa

In West and Central Africa, virulent Newcastle disease virus (NDV) strains of the recently identified genotypes XIV, XVII, and XVIII are enzootic in poultry, representing a considerable threat to the sector. The increasing number of reports of virulent strains in wild birds at least in other parts of the world raised the question of a potential role of wild birds in the spread of virulent NDV in sub-Saharan Africa as well. We investigated 1,723 asymptomatic birds sampled at live-bird markets and sites important for wild-bird conservation in Nigeria and 19 sick or dead wild birds in Côte d''Ivoire for NDV class I and II. Typical avirulent wild-type genotype I strains were found in wild waterfowl in wetlands in northeastern Nigeria. They were unrelated to vaccine strains, and the involvement of inter- or intracontinental migratory birds in their circulation in the region is suggested. Phylogenetic analyses also revealed that genotype VI strains found in pigeons, including some putative new subgenotype VIh and VIi strains, were introduced on multiple separate occasions in Nigeria. A single virulent genotype XVIII strain was found in a dead wild bird in Côte d''Ivoire, probably as a result of spillover from sick poultry. In conclusion, screening of wild birds and pigeons for NDV revealed the presence a variety of virulent and avirulent strains in West Africa but did not provide strong evidence that wild birds play an important role in the spread of virulent strains in the region.     

Next-Generation Sequencing Reveals Frequent Opportunities for Exposure to Hepatitis C Virus in Ghana

Globally, hepatitis C Virus (HCV) infection is responsible for a large proportion of persons with liver disease, including cancer. The infection is highly prevalent in sub-Saharan Africa. West Africa was identified as a geographic origin of two HCV genotypes. However, little is known about the genetic composition of HCV populations in many countries of the region. Using conventional and next-generation sequencing (NGS), we identified and genetically characterized 65 HCV strains circulating among HCV-positive blood donors in Kumasi, Ghana. Phylogenetic analysis using consensus sequences derived from 3 genomic regions of the HCV genome, 5''-untranslated region, hypervariable region 1 (HVR1) and NS5B gene, consistently classified the HCV variants (n = 65) into genotypes 1 (HCV-1, 15%) and genotype 2 (HCV-2, 85%). The Ghanaian and West African HCV-2 NS5B sequences were found completely intermixed in the phylogenetic tree, indicating a substantial genetic heterogeneity of HCV-2 in Ghana. Analysis of HVR1 sequences from intra-host HCV variants obtained by NGS showed that three donors were infected with >1 HCV strain, including infections with 2 genotypes. Two other donors share an HCV strain, indicating HCV transmission between them. The HCV-2 strain sampled from one donor was replaced with another HCV-2 strain after only 2 months of observation, indicating rapid strain switching. Bayesian analysis estimated that the HCV-2 strains in Ghana were expanding since the 16^th century. The blood donors in Kumasi, Ghana, are infected with a very heterogeneous HCV population of HCV-1 and HCV-2, with HCV-2 being prevalent. The detection of three cases of co- or super-infections and transmission linkage between 2 cases suggests frequent opportunities for HCV exposure among the blood donors and is consistent with the reported high HCV prevalence. The conditions for effective HCV-2 transmission existed for ~ 3–4 centuries, indicating a long epidemic history of HCV-2 in Ghana.     

Nucleotide sequence variation of the envelope protein gene identifies two distinct genotypes of yellow fever virus.

The evolution of yellow fever virus over 67 years was investigated by comparing the nucleotide sequences of the envelope (E) protein genes of 20 viruses isolated in Africa, the Caribbean, and South America. Uniformly weighted parsimony algorithm analysis defined two major evolutionary yellow fever virus lineages designated E genotypes I and II. E genotype I contained viruses isolated from East and Central Africa. E genotype II viruses were divided into two sublineages: IIA viruses from West Africa and IIB viruses from America, except for a 1979 virus isolated from Trinidad (TRINID79A). Unique signature patterns were identified at 111 nucleotide and 12 amino acid positions within the yellow fever virus E gene by signature pattern analysis. Yellow fever viruses from East and Central Africa contained unique signatures at 60 nucleotide and five amino acid positions, those from West Africa contained unique signatures at 25 nucleotide and two amino acid positions, and viruses from America contained such signatures at 30 nucleotide and five amino acid positions in the E gene. The dissemination of yellow fever viruses from Africa to the Americas is supported by the close genetic relatedness of genotype IIA and IIB viruses and genetic evidence of a possible second introduction of yellow fever virus from West Africa, as illustrated by the TRINID79A virus isolate. The E protein genes of American IIB yellow fever viruses had higher frequencies of amino acid substitutions than did genes of yellow fever viruses of genotypes I and IIA on the basis of comparisons with a consensus amino acid sequence for the yellow fever E gene. The great variation in the E proteins of American yellow fever virus probably results from positive selection imposed by virus interaction with different species of mosquitoes or nonhuman primates in the Americas.     

HCV基因型的差异性流行与进化

丙型肝炎病毒(Hepatitis C virus,HCV)是导致慢性肝炎的主要病原体之一,全球感染人数大约为1.7亿。HCV基因组具有高度变异特性,利用现代遗传分类方法,可将HCV分为6个基因型和80多个基因亚型。不同HCV基因型、亚型的分布与流行具有明显地域特性:1型、2型呈全球流行态势,3型主要流行于亚洲、北美及欧洲部分地区,4型主要流行于中非、中东和欧洲地区,5型主要发现于非洲和欧洲部分国家,6型则主要在东南亚和北美地区流行。我国流行的HCV有1、2、3和6四种基因型,北方仍以1b和2a型为主要流行基因型,近年来3型和6型在华南、西南地区快速传播。据推断,云南将可能成为我国HCV流行与传播的重要源头,引起目前HCV基因型/亚型分布的较大变化,并呈现多样化的传播方式。通过溯祖理论和进化分子钟等分析方法,了解HCV不同基因型差异性流行与进化,对研究HCV的分子流行病学特征,对应性制定丙型肝炎的预防控制策略具有重要意义。     

Investigating the origin and spread of hepatitis C virus genotype 5a

Epidemiological and phylogenetic studies of hepatitis C virus (HCV) have identified six major HCV genotypes and have attempted to characterize their origin and spread worldwide. Putative regions of endemic infection have been identified for all HCV genotypes except HCV genotype 5a. Although HCV genotype 5a was previously thought to be largely restricted to the northern part of South Africa, this study reports an unexpected cluster of the genotype in West Flanders Province in Belgium. To investigate the molecular epidemiology of this cluster and of HCV genotype 5a in general, a rigorous phylogenetic analysis of Belgian and South African HCV genotype 5a samples was performed. Remarkably, the Belgian and South African strains form two distinct clusters of similar diversity. We used a Bayesian coalescent method to estimate the rate of virus spread through time for HCV genotype 5a in both regions. Our results indicate that HCV genotype 5a strains have been spreading independently in Belgium and South Africa for more than 100 years, with a rate of spread characteristic of an epidemic genotype. These findings have major implications for tracing the origin of HCV genotype 5a. Here, we speculate about the possible origins of these clusters.     

黄热病现状及其疫苗的研究与应用

黄热病(yellow fever,YF)是一种经由伊蚊叮咬传播的急性出血性传染病,流行区主要集中在非洲西部、南美洲等热带地区,临床症状包括高热、恶心、呕吐、黄疸、出血等。黄热病的病原体为黄热病毒(yellow fever virus, YFV),是黄病毒科黄病毒属的单股正链RNA病毒。目前,YF治疗尚无特效药物,以对症治疗和支持治疗为主,接种YF-17D减毒活疫苗是最有效的预防方法。尽管YF-17D已被全球认可,但是疫苗接种所致的不良反应时有报道。本文对近年黄热病的流行情况、疫苗研究进展和应用进行综述。     

Genotyping Mycobacterium ulcerans and Mycobacterium marinum by using mycobacterial interspersed repetitive units

A novel category of variable tandem repeats (VNTR) called mycobacterial interspersed repetitive units (MIRUs) has been identified for Mycobacterium ulcerans (n = 39), M. marinum (n = 27), and one related organism. Fifteen MIRU loci were identified in the genome of M. marinum and were used to genotype M. ulcerans, M. marinum, and an M. marinum-like organism that is considered a possible missing link between M. marinum and M. ulcerans. Seven MIRU loci were polymorphic, and locus-specific PCRs for four of these loci differentiated seven M. ulcerans genotypes, four M. marinum genotypes, and a unique genotype for the missing link organism. The seven M. ulcerans genotypes were related to six different geographic origins of isolates. All isolates from West and Central Africa, including old and recent isolates, belonged to the same genotype, emphasizing the great spatiotemporal homogeneity among African isolates. Unlike the M. ulcerans genotypes, the four M. marinum genotypes could not be clearly related to the geographic origins of the isolates. According to MIRU-VNTR typing, all M. ulcerans and M. marinum isolates of American origin were closely related, suggesting a common American ancestor for these two pathogenic species on the American continents. MIRU typing has significant potential value for discriminating between reoccurrence and reinfection for M. ulcerans disease.     

Additional haplogroups of Toxoplasma gondii out of Africa: population structure and mouse-virulence of strains from Gabon

Background

Toxoplasma gondii is found worldwide, but distribution of its genotypes as well as clinical expression of human toxoplasmosis varies across the continents. Several studies in Europe, North America and South America argued for a role of genotypes in the clinical expression of human toxoplasmosis. Genetic data concerning T. gondii isolates from Africa are scarce and not sufficient to investigate the population structure, a fundamental analysis for a better understanding of distribution, circulation, and transmission.

Methodology/Principal Findings

Seropositive animals originating from urban and rural areas in Gabon were analyzed for T. gondii isolation and genotyping. Sixty-eight isolates, including one mixed infection (69 strains), were obtained by bioassay in mice. Genotyping was performed using length polymorphism of 13 microsatellite markers located on 10 different chromosomes. Results were analyzed in terms of population structure by Bayesian statistical modeling, Neighbor-joining trees reconstruction based on genetic distances, F _ST and linkage disequilibrium. A moderate genetic diversity was detected. Three haplogroups and one single genotype clustered 27 genotypes. The majority of strains belonged to one haplogroup corresponding to the worldwide Type III. The remaining strains were distributed into two haplogroups (Africa 1 and 3) and one single genotype. Mouse virulence at isolation was significantly different between haplogroups. Africa 1 haplogroup was the most virulent.

Conclusion

Africa 1 and 3 haplogroups were proposed as being new major haplogroups of T. gondii circulating in Africa. A possible link with strains circulating in South and Central America is discussed. Analysis of population structure demonstrated a local spread within a rural area and strain circulation between the main cities of the country. This circulation, favored by human activity could lead to genetic exchanges. For the first time, key epidemiological questions were addressed for the West African T. gondii population, using the high discriminatory power of microsatellite markers, thus creating a basis for further epidemiological and clinical investigations.     

Pathogenic variability of downy mildew (Sclerospora graminicola) on pearl millet. I. Host cultivar reactions to infection by different pathogen isolates

Cultivars of pearl millet were challenged by isolates of downy mildew collected from various locations in West Africa and India in order to ascertain whether variability in cultivar response was genetically or environmentally determined. Results of experiments, in Polythene tunnels which imitated tropical field conditions, were confirmed by more precise experiments in an isolation plant propagator. The most important conclusion was that variation is determined by host and pathogen genotypes. West African isolates of the pathogen were generally more pathogenic than Indian isolates. However, there were also substantial differences between two isolates collected from different host cultivars at the same location in Upper Volta. Cultivar ICH105 differentiated between West African and Indian isolates. Cultivars 700516 and MBH110 also showed differential responses between isolates. In contrast two distinct types of symptom expression were recorded and found to be characteristic of cultivar genotype, independent of pathogen isolate. The possibility that both race specific and race non-specific resistance may coexist in this little understood pathosystem is discussed and the practical implications are considered.     

Epidemiology and molecular characterization of hepatitis B virus in Luanda, Angola

An estimated 360 million people are infected with hepatitis B virus (HBV) worldwide. Among these, 65 million live in Africa. Despite the high levels of hepatitis B in Africa, HBV epidemiology is still poorly documented in most African countries. In this work, the epidemiological and molecular characteristics of HBV infection were evaluated among the staff, visitors and adult patients (n = 508) of a public hospital in Luanda, Angola. The overall prevalence of hepatitis B core antibody (anti-HBc) and hepatitis B surface antigen was 79.7% and 15.1%, respectively. HBV infection was higher in males and was more prevalent in individuals younger than 50 years old. HBV-DNA was detected in 100% of HBV "e" antigen-positive serum samples and in 49% of anti-hepatitis Be antibody-positive samples. Thirty-five out of the 40 HBV genotypes belonged to genotype E. Circulation of genotypes A (4 samples) and D (1 sample) was also observed. The present study demonstrates that HBV infection is endemic in Luanda, which has a predominance of genotype E. This genotype is only sporadically found outside of Africa and is thought to have emerged in Africa at a time when the trans-Atlantic slave trade had stopped.     

A hotspot of Toxoplasma gondii Africa 1 lineage in Benin: How new genotypes from West Africa contribute to understand the parasite genetic diversity worldwide

Through international trades, Europe, Africa and South America share a long history of exchanges, potentially of pathogens. We used the worldwide parasite Toxoplasma gondii to test the hypothesis of a historical influence on pathogen genetic diversity in Benin, a West African country with a longstanding sea trade history. In Africa, T. gondii spatial structure is still non-uniformly studied and very few articles have reported strain genetic diversity in fauna and clinical forms of human toxoplasmosis so far, even in African diaspora. Sera from 758 domestic animals (mainly poultry) in two coastal areas (Cotonou and Ouidah) and two inland areas (Parakou and Natitingou) were tested for T. gondii antibodies using a Modified Agglutination Test (MAT). The hearts and brains of 69 seropositive animals were collected for parasite isolation in a mouse bioassay. Forty-five strains were obtained and 39 genotypes could be described via 15-microsatellite genotyping, with a predominance of the autochthonous African lineage Africa 1 (36/39). The remaining genotypes were Africa 4 variant TUB2 (1/39) and two identical isolates (clone) of Type III (2/39). No difference in terms of genotype distribution between inland and coastal sampling sites was found. In particular, contrarily to what has been described in Senegal, no type II (mostly present in Europe) was isolated in poultry from coastal cities. This result seems to refute a possible role of European maritime trade in Benin despite it was one of the most important hubs during the slave trade period. However, the presence of the Africa 1 genotype in Brazil, predominant in Benin, and genetic analyses suggest that the triangular trade was a route for the intercontinental dissemination of genetic strains from Africa to South America. This supports the possibility of contamination in humans and animals with potentially imported virulent strains.     

Emergence of dengue virus serotype 2 in Mauritania and molecular characterization of its circulation in West Africa

The number of sporadic and epidemic dengue fever cases have reportedly been increasing in recent years in some West African countries, such as Senegal and Mali. The first epidemic of laboratory-confirmed dengue occurred in Nouakchott, the capital city of Mauritania situated in the Saharan desert, in 2014. On-site diagnosis of dengue fever was established using a rapid diagnostic test for dengue. In parallel, the presence of Aedes aegypti mosquitoes in the city was confirmed. The initial diagnosis was confirmed by RT-PCR, which showed that all samples from the 2014 dengue epidemic in Nouakchott were dengue virus serotype 2 (DENV-2). The whole genome or envelope protein gene of these strains, together with other DENV-2 strains obtained from travelers returning from West African countries to France between 2016 and 2019 (including two Mauritanian strains in 2017 and 2018), were sequenced. Phylogenetic analysis suggested a recent emergence of an epidemic strain from the cosmopolitan genotype belonging to West African cosmopolitan lineage II, which is genetically distinct from African sylvatic genotype. The origin of this DENV-2 lineage is still unknown, but our data seem to suggest a recent and rapid dispersion of the epidemic strain throughout the region. More complete genome sequences of West African DENV-2 are required for a better understanding of the dynamics of its circulation. Arboviral surveillance and outbreak forecasting are urgently needed in West Africa.     

Polyomavirus JC genotypes in an urban United States population reflect the history of African origin and genetic admixture in modern African Americans

The human polyomavirus JC (JC virus), a small, circular, double-stranded DNA virus, has a worldwide distribution and is excreted harmlessly in urine by 20% to 70% of adults. DNA sequence analysis has identified seven distinct genotypes that likely coevolved with modern humans, although the mode of virus transmission is unknown. Type 1 is European in its distribution. Types 2 and 7 are Asian, while Types 3 and 6 are African. Type 4, closely related to Type 1, is of uncertain origin, having been found in population groups in parts of Europe and in the United States, but not in Africa. Here we have studied the JCV partial genomic DNA sequences amplified by polymerase chain reaction techniques from urines of an urban, mainly African American population cohort from Washington, D.C. The predominant genotype identified was Type 4 (32/78 JCV strains, 41%). Type 1 strain was found in 32% of African Americans, while JCV Type 3 strain was found in 18% of African Americans. These African strains have persisted in modern African Americans after 200 to 400 years of minority existence and genetic admixture in the New World. An ancient West African genotype, Type 6, was absent in this African American cohort. However, one Type 6 strain was found in a patient from Sierra Leone (West Africa), domiciled in the United States for 20 years. Type 2A, the most common subtype in Native Americans, was seen in only two African-Americans (3%). A Type 7 strain, previously reported only in Taiwan and South China, was identified in a Vietnamese immigrant. These data support the history of African origin, migration, and genetic admixture of modern African Americans. Analysis of JCV strains in the present American populations provides a novel tool for reconstructing human migrations and genetic admixture in the New World.     

Prevalence of HBV genotypes among Egyptian hepatitis patients

Phylogenetic analysis has led to the classification of hepatitis B virus into eight genotypes, designated A to H. The genotypes have differences in biological properties and show heterogeneity in their global distribution. These attributes of the genotypes may account not only for differences in the prevalence of hepatitis B virus mutants in various geographic regions, but also makes them responsible for differences in the clinical outcome and response to antiviral treatment in different population groups. Africa is one of the highly endemic regions of HBV with five genotypes (A–E) identified. Almost all patients in the Mediterranean area are infected with genotype D. However, there is little information of genotype distribution in Egypt. A total of 140 Egyptian patients with hepatitis B surface antigen (HBsAg) positive were enrolled in this study. Of the 140 patients, only 100 patients were HBV DNA positive and only these were included in the study. They were classified into 20 patients with acute hepatitis (AH), 75 patients with chronic active hepatitis (CAH) and 5 patients with hepatocellular carcinoma (HCC). HBV genotypes were determined using INNO-LiPA methodology which is based on the reversed hybridization principle. This study showed that genotype D constituted 87% of the total infections (75 CAH cases, 7 AH cases and 5 HCC cases). The other 13% showed mixed infections of D/F. These findings show that the most prevalent genotype in Egypt is genotype D especially in CAH and HCC patients while the mixed type D/F is only encountered in AH.     

Host-cell interaction of attenuated and wild-type strains of yellow fever virus can be differentiated at early stages of hepatocyte infection

Yellow fever (YF) virus is currently found in tropical Africa and South America, and is responsible for a febrile to severe illness characterized by organ failure and shock. The attenuated YF 17D strain, used in YF vaccine, was derived from the wild-type strain Asibi. Although studies have been done on genetic markers of YF virulence, differentiation of the two strains in terms of host-cell interaction during infection remains elusive. As YF wild-type strains are hepatotropic, we chose a hepatic cell line (HepG2) to study YF virus-host cell interaction. HepG2 cells rapidly produced high titres of infectious viral particles for 17D and Asibi YF strains. However, HepG2 cells were more susceptible to the attenuated 17D virus infection, and only this virus strain induced early apoptosis in these cells. Molecular markers specific for the 17D virus were identified by microarray analysis and confirmed by quantitative RT-PCR analysis. As early as 1h postinfection, three genes, (IEX-1, IRF-1, DEC-1) all implicated in apoptosis pathways, were upregulated. Later in infection (48 h) two other genes (HSP70-1A and 1B), expressed in cases of cellular stress, were highly upregulated in 17D-infected HepG2 cells. The early specific upregulation of these cellular genes in HepG2 cells may be considered markers of the 17D virus. This study on the YF attenuated strain gives a new approach to the analysis of the factors involved in virus attenuation.     

Technical viability of the YF MAC-HD ELISA kit for use in yellow fever-endemic regions

Yellow fever (YF), an arboviral disease, affects an estimated 200,000 people and causes 30,000 deaths per year and recently has caused major epidemics in Africa and South America. Timely and accurate diagnosis of YF is critical for managing outbreaks and implementing vaccination campaigns. A YF immunoglobulin M (IgM) antibody-capture (MAC) enzyme-linked immunosorbent assay (ELISA) kit, the YF MAC-HD, was successfully introduced starting in 2018 to laboratories in Africa and South America. The YF MAC-HD kit can be performed in 3.5 hours, test up to 24 samples, and includes all reagents necessary to perform the test, except for water used to dilute wash buffer. In 2018 and 2019, a total of 56 laboratory personnel from 39 countries in Africa and South America were trained to use the kit during workshops, followed by take-home YF IgM proficiency testing (PT) exercises. Participants received either a 10- or 20-sample YF PT panel and performed testing using the YF MAC-HD kit. All countries obtained 90% or higher correct results. These results verified the technical viability and transferability of YF MAC-HD kit use for laboratories in YF-endemic countries.     

Molecular epidemiology of anthrax cases associated with recreational use of animal hides and yarn in the United States

To determine potential links between the clinical isolate to animal products and their geographic origin, we genotyped (MLVA-8, MVLA-15, and canSNP analysis) 80 environmental and 12 clinical isolates and 2 clinical specimens from five cases of anthrax (California in 1976 [n = 1], New York in 2006 [n = 1], Connecticut in 2007 [n = 2], and New Hampshire in 2009[n = 1]) resulting from recreational handling of animal products. For the California case, four clinical isolates were identified as MLVA-8 genotype (GT) 76 and in the canSNP A.Br.Vollum lineage, which is consistent with the Pakistani origin of the yarn. Twenty eight of the California isolates were in the A.Br.Vollum canSNP lineage and one isolate was in the A.Br. 003/004 canSNP sub-group. All 52 isolates and both clinical specimens related to the New York and Connecticut cases were MLVA-8 GT 1. The animal products associated with the NY and CT cases were believed to originate from West Africa, but no isolates from this region are available to be genotyped for comparison. All isolates associated with the New Hampshire case were identical and had a new genotype (GT 149). Isolates from the NY, CT and NH cases diverge from the established canSNP phylogeny near the base of the A.Br.011/009. This report illustrates the power of the current genotyping methods and the dramatically different epidemiological conditions that can lead to infections (i.e., contamination by a single genotype versus widespread contamination of numerous genotypes). These cases illustrate the need to acquire and genotype global isolates so that accurate assignments can be made about isolate origins.     

Genetic Characterization of Toxoplasma gondii Isolates and Toxoplasmosis Seroprevalence in Stray Cats of ?zmir,Turkey

Currently, some Toxoplasma gondii genotypes are being associated with serious clinical presentations. A recent report showing the Africa 1 genotype in two local congenital toxoplasmosis cases acquired in Turkey formed the basis of this study because atypical Africa 1 genotype is most frequently detected in animals and patients from sub-Saharan Africa. Since stray cats are considered as the linkage between wild life and urban life in T. gondii transmission, the present study aimed to isolate and characterize T. gondii strains circulating in stray cats of İzmir (Western Turkey). A secondary objective was to determine toxoplasmosis seroprevalence in this cat population. Tissues obtained from 100 deceased stray cats were bioassayed and isolated strains were genotyped using 15 microsatellite markers. In addition, toxoplasmosis seroprevalence was analyzed in 1121 cat sera collected from several large veterinary clinics in İzmir. Among the 22 isolates, 19 were Type II (86.3%), two were Type III (9%) and one was Africa 1 genotype (4.5%). The overall seropositivity rates in cats were 42–48% and 33.4–34.4% according to IFA and ELISA, respectively. Seroprevalence in deceased cats was significantly higher than in healthy cats (P = 0.0033). Finding both the major clonal Type II lineage together with the Type III lineage also found in Middle East, and an atypical genotype, Africa 1 appears consistent with the specific geographic location of Turkey between three continents and raises the possibility of transportation of these strains between continents through trade routes or long distance migratory birds. In addition, the first large study of toxoplasma seroprevalence in a stray cat population was also reported. The relatively high seropositivity rates and the variety of T. gondii genotypes confirm the local stray cat population as a risk factor for human toxoplasmosis in İzmir.