American Strains Research Articles

Temporal trends of cutaneo-mucous histoplasmosis in persons living with HIV in French Guiana: Early diagnosis defuses South American strain dermotropism.

Histoplasmosis is the most frequent opportunistic infection and the first cause of mortality in HIV-infected patients in French Guiana and presumably in much of Latin America. Mucocutaneous lesions of histoplasmosis are considered as rare and late manifestations of the disease. It has been debated whether the greater proportion of cutaneo-mucous presentations in South America relative to the USA was the reflection of Histoplasma strains with increased dermotropism or simply delayed diagnosis and advanced immunosuppression. The objective of this study was to describe the clinical presentation, frequency, prognosis and temporal trends of cutaneomucous histoplasmosis in French Guiana. A retrospective study of patients with AIDS-related disseminated histoplasmosis followed in the three hospitals of French Guiana was performed between 1981 and 2014. Incident cases of histoplasmosis, proved by pathology and/or mycological examinations, were studied. Mucocutaneous histoplasmosis was confirmed by a positive cutaneous or mucosal biopsy. Mucocutaneous lesions were polymorphic. Ninety percent of patients were profoundly immunocompromised patients (CD4<50/mm3) and over 80% were not on antiretroviral treatment. The frequency of mucocutaneous forms and case fatality of disseminated histoplasmosis within one month of antifungal treatment significantly decreased over time (p<0,001). In this South American territory, diagnostic and therapeutic improvements have led to the quasi disappearance of cutaneous manifestations. There may be South American dermotropism in the laboratory but at the bedside early diagnosis seems to be the main parameter explaining the proportion of cutaneomucous presentations in South America relative to the USA.

Outbreak of a novel disease caused by Staphylococcus pseudintermedius in raccoon dogs (Nyctereutes procyonoides).

This study reports outbreak of a new disease caused by Staphylococcus pseudintermedius (S.pseudintermedius) in raccoon dogs. The disease occurred in a breeding farm of raccoon dogs in Guan County of Shandong Province in China in August of 2019. 47% (425/896) of the raccoon dogs showed some abnormal symptoms; 17.6% (75/425) of which had severe skin and soft tissue infections (SSTIs), dyspnoea and severe pathological lesions in lungs, livers, etc; and 4.2% (18/425) of which died within 4weeks. The pathogen of the disease was identified as S. pseudintermedius by mass spectrometer detection, animal pathogenicity tests, microscopic examination and biochemical reaction tests. Its nucleotide homology of 16S rRNA gene was 100% with that of other published strains, and its genotype was between the American and Brazilian strains from other animals. The isolated S.pseudintermedius strain from the diseased raccoon dogs could cause ulceration and suppuration in the skins and severe pathological lesions not only in raccoon dogs, but also in mice; and it is confirmed as a methicillin-resistant S.pseudintermedius (MRSP) strain by the amplification of mecA gene; and 12 sensitive drugs were screened by drug sensitivity tests. Full attention should be paid to the great economic loss and the potential zoonotic risk caused by the S.pseudintermedius in raccoon dogs, and this study can provide a reference for the clinical diagnosis and treatment of this new disease.

Global and regional dispersal patterns of hepatitis B virus genotype E from and in Africa: A full-genome molecular analysis.

Description of the spatial characteristics of viral dispersal is important in understanding the history of infections. Nine hepatitis B virus (HBV) genotypes (A-I), and a putative 10th genotype (J), with distinct geographical distribution, are recognized. In sub-Saharan Africa (sub)-genotypes A1, D3 and E circulate, with E predominating in western Africa (WA), where HBV is hyperendemic. The low genetic diversity of genotype E (HBV/E) suggests its recent emergence. Our aim was to study the dispersal of HBV/E using full-length, non-redundant and non-recombinant sequences available in public databases. HBV/E was confirmed, and the phylogeny reconstruction performed using maximum likelihood (ML) with bootstrapping. Phylogeographic analysis was conducted by reconstruction of ancestral states using the criterion of parsimony on the estimated ML phylogeny. 46.5% of HBV/E sequences were found within monophyletic clusters. Country-wise analysis revealed the existence of 50 regional clusters. Sequences from WA were located close to the root of the tree, indicating this region as the most probable origin of the HBV/E epidemic and expanded to other geographical regions, within and outside of Africa. A localized dispersal was observed with sequences from Nigeria and Guinea as compared to other WA countries. Based on the sequences available in the databases, the phylogenetic results suggest that European strains originated primarily from WA whereas a majority of American strains originated in Western Central Africa. The differences in regional dispersal patterns of HBV/E suggest limited cross-border transmissions because of restricted population movements.

Borrelia miyamotoi strain LB-2001 retains plasmids and infectious phenotype throughout continuous culture passages as evaluated by multiplex PCR

Borrelia miyamotoi is a tick-borne spirochete of the relapsing fever borrelia group and an emerging pathogen of public health significance. The genomes of relapsing fever borreliae and Lyme disease borreliae consist of multiple linear and circular plasmids in addition to the chromosome. Previous work with B. burgdorferi sensu lato found diminished infectivity upon continuous in vitro culture passage that was attributable to plasmid loss. The effect of long-term culture passage on B. miyamotoi is not known. We generated a series of plasmid-specific primer sets and developed a multiplex PCR assay to detect the 14 known plasmids of B. miyamotoi North American strains LB-2001 and CT13−2396. We assessed the plasmid content of B. miyamotoi LB-2001 over 64 culture passages spanning 15 months and determined that strain LB-2001 retained all plasmids upon prolonged in vitro cultivation and remained infectious in mice. We also found that strain LB-2001 lacks plasmid lp20−1 which is present in strain CT13−2396. These results suggest that B. miyamotoi remains genetically stable when cultured and passaged in vitro.

Phylogenomic analysis of Mycoplasma bovis from Belgian veal, dairy and beef herds

M. bovis is one of the leading causes of respiratory disease and antimicrobial use in cattle. The pathogen is widespread in different cattle industries worldwide, but highest prevalence is found in the veal industry. Knowledge on M. bovis strain distribution over the dairy, beef and veal industries is crucial for the design of effective control and prevention programs, but currently undocumented. Therefore, the present study evaluated the molecular epidemiology and genetic relatedness of M. bovis isolates obtained from Belgian beef, dairy and veal farms, and how these relate to M. bovis strains obtained worldwide. Full genomes of one hundred Belgian M. bovis isolates collected over a 5-year period (2014–2019), obtained from 27 dairy, 38 beef and 29 veal farms, were sequenced by long-read nanopore sequencing. Consensus sequences were used to generate a phylogenetic tree in order to associate genetic clusters with cattle sector, geographical area and year of isolation. The phylogenetic analysis of the Belgian M. bovis isolates resulted in 5 major clusters and 1 outlier. No sector-specific M. bovis clustering was identified. On a world scale, Belgian isolates clustered with Israeli, European and American strains. Different M. bovis clusters circulated for at least 1.5 consecutive years throughout the country, affecting all observed industries. Therefore, the high prevalence in the veal industry is more likely the consequence of frequent purchase from the dairy and beef industry, than that a reservoir of veal specific strains on farm would exist. These results emphasize the importance of biosecurity in M. bovis control and prevention.

Phylogenomic Analysis Supports Two Possible Origins for Latin American Strains of Vibrio parahaemolyticus Associated with Acute Hepatopancreatic Necrosis Disease (AHPND).

Acute hepatopancreatic necrosis disease (AHPND) is a severe disease affecting recently stocked cultured shrimps. The disease is mainly caused by V. parahaemolyticus that harbors the pVA1 plasmid; this plasmid contains the pirA and pirB genes, which encode a delta-endotoxin. AHPND originated in China in 2009 and has since spread to several other Asian countries and recently to Latin America (2013). Many Asian strains have been sequenced, and their sequences are publicly accessible in scientific databases, but only four strains from Latin America have been reported. In this study, we analyzed nine pVA1-harboring V. parahaemolyticus sequences from strains isolated in Mexico along with the 38 previously available pVA1-harboring V. parahaemolyticus sequences and the reference strain RIMD 2210633. The studied sequences were clustered into three phylogenetic clades (Latin American, Malaysian, and Cosmopolitan) through pangenomic and phylogenomic analysis. The nucleotide sequence alignment of the pVA1 plasmids harbored by the Asian and Latin American strains confirmed that the main structural difference in the plasmid between the Asian and Latin American strains is the absence of the Tn3 transposon in the Asian strains; in addition, some deletions in the pirAB region were found in two of the Latin American strains. Our study represents the most robust and inclusive phylogenomic analysis of pVA1-harboring V. parahaemolyticus conducted to date and provides insight into the epidemiology of AHPND. In addition, this study highlights that disease diagnosis through the detection of the pirA and pirB genes is an inadequate approach due to the instability of these genes.

Circulation of Indigenous Bovine Respiratory Syncytial Virus Strains in Turkish Cattle: The First Isolation and Molecular Characterization.

Simple SummaryBovine respiratory syncytial virus (BRSV) is an important pathogen of both dairy and beef cattle, and causes huge economic losses annually across the world. This study reports the identification, isolation, and molecular characterization of a new BRSV (subgroup III) strain collected from respiratory distressed cattle in Turkey. The three field isolates obtained showed 100% similarity to each other at the nucleotide (nt) level and were found to be 99.49% and 99.22% identical to another Turkish strain—KY499619—at both (nt) and amino acid (aa) levels, respectively. They were also 97.43% (nt) and 98.44% (aa) similar to the American reference strain KU159366. This important information will inform Turkish BRSV diagnostic and control strategies, as well as highlight the urgent need to better understand the burden that BRSV is placing on the Turkish agricultural sector.Bovine respiratory disease (BRD) is a huge economic burden on the livestock industries of countries worldwide. Bovine respiratory syncytial virus (BRSV) is one of the most important pathogens that contributes to BRD. In this study, we report the identification and first isolation, with molecular characterization, of a new BRSV strain from lung specimens of three beef cows in Turkey that died from respiratory distress. After the screening of lung tissues for BRD-associated viruses using a multiscreen antigen-ELISA, a BRSV antigen was detected. This was then confirmed by real-time RT-PCR specific for BRSV. Following confirmation, virus isolation was conducted in MDBK cell cultures and clear CPE, including syncytia compatible with BRSV, were detected. RT-nested PCR, using F gene-specific primers, was performed on the cultured isolates, and the products were sequenced and deposited to Genbank with accession numbers MT179304, MT024766, and MT0244767. Phylogenetic analysis of these sequences indicated that the cattle were infected with BRSV from subgroup III and were closely related to previously identified American and Turkish strains, but contained some amino acid and nucleotide differences. This research paves the way for further studies on the molecular characteristics of natural BRSV isolates, including full genome analysis and disease pathogenesis, and also contributes to the development of robust national strategies against this virus.

A 500-year tale of co-evolution, adaptation, and virulence: Helicobacter pylori in the Americas

Helicobacter pylori is a common component of the human stomach microbiota, possibly dating back to the speciation of Homo sapiens. A history of pathogen evolution in allopatry has led to the development of genetically distinct H. pylori subpopulations, associated with different human populations, and more recent admixture among H. pylori subpopulations can provide information about human migrations. However, little is known about the degree to which some H. pylori genes are conserved in the face of admixture, potentially indicating host adaptation, or how virulence genes spread among different populations. We analyzed H. pylori genomes from 14 countries in the Americas, strains from the Iberian Peninsula, and public genomes from Europe, Africa, and Asia, to investigate how admixture varies across different regions and gene families. Whole-genome analyses of 723 H. pylori strains from around the world showed evidence of frequent admixture in the American strains with a complex mosaic of contributions from H. pylori populations originating in the Americas as well as other continents. Despite the complex admixture, distinctive genomic fingerprints were identified for each region, revealing novel American H. pylori subpopulations. A pan-genome Fst analysis showed that variation in virulence genes had the strongest fixation in America, compared with non-American populations, and that much of the variation constituted non-synonymous substitutions in functional domains. Network analyses suggest that these virulence genes have followed unique evolutionary paths in the American populations, spreading into different genetic backgrounds, potentially contributing to the high risk of gastric cancer in the region.

Antifungal Activities of Ziziphus mauritiana against Candida albicans: In Vitro Study

Medicinal plants have been discovered and used in traditional medicine and pharmaceutical industries since centuries. In the current study, Ziziphus mauritiana leaves was used as it is rich with many biological active compounds such as flavonoids, polyphenols, sapronins and tanins. Previous studies reported the antibacterial and antifungal effects of Z. mauritiana towards various microorganisms. However, the antifungal activities of Z. mauritiana methanol extracts on Candida albicans (Clinical and American Type Culture Collection Strains) have not been discovered extensively.Objectives: Therefore, the aim of the current study is to investigate the antifungal activity of Z. mauritiana leaves methanol extracts against C. albicans ATCC strain and clinical isolate (from oral cancer patient).Methods: Antifungal susceptibility test (AST) was performed using disc diffusion, minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) to determine the antifungal activity of methanol leaves extract of Ziziphus mauritiana against C. albicans ATCC and clinical isolates.Results: The results obtained showed that there is no zone of inhibition seen from disk diffusion test for both strains. However, the minimal inhibitory and minimal fungicidal concentration showed that Ziziphus mauritiana methanol extracts was able to inhibit C. albicans clinical isolate but not ATCC strain at 500mg/mL.Conclusion: The finding of this study suggests that that Ziziphus mauritiana leaves methanol extract showed promising results against Candida albicans. Thus, it can be used as a source for functional ingredients for pharmaceutical drug industries in-order to reduce or inhibit oral fungal infection.

Hypervariable Domain of nsP3 of Eastern Equine Encephalitis Virus Is a Critical Determinant of Viral Virulence.

Eastern equine encephalitis virus (EEEV) is the most pathogenic member of the Alphavirus genus in the Togaviridae family. This virus continues to circulate in the New World and has a potential for deliberate use as a bioweapon. Despite the public health threat, to date no attenuated EEEV variants have been applied as live EEEV vaccines. Our previous studies demonstrated the critical function of the hypervariable domain (HVD) in EEEV nsP3 for the assembly of viral replication complexes (vRCs). EEEV HVD contains short linear motifs that recruit host proteins required for vRC formation and function. In this study, we developed a set of EEEV mutants that contained combinations of deletions in nsP3 HVD and clustered mutations in capsid protein, and tested the effects of these modifications on EEEV infection in vivo These mutations had cumulative negative effects on viral ability to induce meningoencephalitis. The deletions of two critical motifs, which interact with the members of cellular FXR and G3BP protein families, made EEEV cease to be neurovirulent. The additional clustered mutations in capsid protein, which affect its ability to induce transcriptional shutoff, diminished EEEV's ability to develop viremia. Most notably, despite the inability to induce detectable disease, the designed EEEV mutants remained highly immunogenic and, after a single dose, protected mice against subsequent infection with wild-type (wt) EEEV. Thus, alterations of interactions of EEEV HVD and likely HVDs of other alphaviruses with host factors represent an important direction for development of highly attenuated viruses that can be applied as live vaccines.IMPORTANCE Hypervariable domains (HVDs) of alphavirus nsP3 proteins recruit host proteins into viral replication complexes. The sets of HVD-binding host factors are specific for each alphavirus, and we have previously identified those specific for EEEV. The results of this study demonstrate that the deletions of the binding sites of the G3BP and FXR protein families in the nsP3 HVD of EEEV make the virus avirulent for mice. Mutations in the nuclear localization signal in EEEV capsid protein have an additional negative effect on viral replication in vivo Despite the inability to cause a detectable disease, the double HVD and triple HVD/capsid mutants induce high levels of neutralizing antibodies. Single immunization protects mice against infection with the highly pathogenic North American strain of EEEV. High safety, the inability to revert to wild-type phenotype, and high immunogenicity make the designed mutants attractive vaccine candidates for EEEV infection.

Application of Bayesian phylogenetic inference modelling for evolutionary genetic analysis and dynamic changes in 2019-nCoV.

The novel coronavirus (2019-nCoV) has recently caused a large-scale outbreak of viral pneumonia both in China and worldwide. In this study, we obtained the entire genome sequence of 777 new coronavirus strains as of 29 February 2020 from a public gene bank. Bioinformatics analysis of these strains indicated that the mutation rate of these new coronaviruses is not high at present, similar to the mutation rate of the severe acute respiratory syndrome (SARS) virus. The similarities of 2019-nCoV and SARS virus suggested that the S and ORF6 proteins shared a low similarity, while the E protein shared the higher similarity. The 2019-nCoV sequence has similar potential phosphorylation sites and glycosylation sites on the surface protein and the ORF1ab polyprotein as the SARS virus; however, there are differences in potential modification sites between the Chinese strain and some American strains. At the same time, we proposed two possible recombination sites for 2019-nCoV. Based on the results of the skyline, we speculate that the activity of the gene population of 2019-nCoV may be before the end of 2019. As the scope of the 2019-nCoV infection further expands, it may produce different adaptive evolutions due to different environments. Finally, evolutionary genetic analysis can be a useful resource for studying the spread and virulence of 2019-nCoV, which are essential aspects of preventive and precise medicine.

Characteristics of the spike and ORF3 genes of porcine epidemic diarrhea virus in Henan and Shanxi provinces of China.

To investigate the epidemic characteristics of porcine epidemic diarrhea virus (PEDV), 135 clinical samples (including intestinal tissues and feces) were collected from diseased piglets during outbreaks of diarrhea from 2015 to 2019 on farms in Henan and Shanxi provinces of China where swine had been immunized with attenuated PEDV (CV777). A total of 86 clinical samples (86/135, 63.7%) were positive for PEDV by RT-PCR, and subsequently, the complete spike (S) and ORF3 genes of 32 PEDV samples were sequenced. Phylogenetic analysis showed that the 32 PEDV strains obtained in this study belonged to group 2 (pandemic variant strains) and had a close relationship to 17 Chinese strains after 2010, two South Korean strains (KNU-1305 and KNU-1807), three American strains (PC22A-P140.BI, USA/Colorado/2013, and USA/OK10240-6/2017) and a Mexican strain (PEDV/MEX/QRO/02/2017), but differed genetically from a South Korean strain (SM98), a European strain (Br1/87), a Chinese strain (LZC), and a vaccine strain (CV777). G2-a subgroup strains were the dominant pandemic variant strains circulating in Henan and Shanxi provinces of China. Furthermore, a cross-recombination event was identified in the S region of the SX/TY2/2017 strain, and the putative parental strains were the epidemic strains CH/GDGZ/2012 and CH/YZ1/2015, identified in China in 2012 and 2015, respectively. These results provide further information about PEDV evolution, which could improve our understanding of the circulation of PEDV in Henan and Shanxi provinces. This information will also be helpful for developing new strategies for prevention and control of variant strains.

Exploring heterologous prime-boost vaccination approaches to enhance influenza control in pigs

Influenza A viruses evolve rapidly to escape host immunity. In swine, this viral evolution has resulted in the emergence of multiple H1 and H3 influenza A virus (IAV) lineages in the United States (US) pig populations. The heterologous prime-boost vaccination strategy is a promising way to deal with diverse IAV infection in multiple animal models. However, whether or not this vaccination strategy is applicable to US swine to impart immunity against infection from North American strains of IAV is still unknown. We performed a vaccination-challenge study to evaluate the protective efficacy of using multivalent inactivated vaccine and/or a live attenuated IAV vaccine (LAIV) in pigs following multiple prime-boost vaccination protocols against a simultaneous H1N1 and H3N2 IAV infection. Our data show that pigs in the heterologous prime-boost vaccination group had more favorable outcomes consistent with a better response against virus challenge than non-vaccinated pigs. Additionally, delivering a multivalent heterologous inactivated vaccine boost to pigs following a single LAIV administration was also beneficial. We concluded the heterologous prime boost vaccination strategy may potentiate responses to suboptimal immunogens and holds the potential applicability to control IAV in the North American swine industry. However, more studies are needed to validate the application of this vaccination approach under field conditions.

Mapping the evolutionary landscape of Zika virus infection in immunocompromised mice

The fundamental basis of how arboviruses evolve in nature and what regulates the adaptive process remain unclear. To address this problem, we established a Zika virus (ZIKV) vector-borne transmission system in immunocompromised mice to study the evolutionary characteristics of ZIKV infection. Using this system, we defined factors that influence the evolutionary landscape of ZIKV infection and show that transmission route and specific organ microenvironments impact viral diversity and defective viral genome production. In addition, we identified in mice the emergence of ZIKV mutants previously seen in natural infections, including variants present in currently circulating Asian and American strains, as well as mutations unique to the mouse infections. With these studies, we have established an insect-to-mouse transmission model to study ZIKV evolution in vivo. We also defined how organ microenvironments and infection route impact the ZIKV evolutionary landscape, providing a deeper understanding of the factors that regulate arbovirus evolution and emergence.

Serologic reactivity of canine sera to Sarcocystis neurona and Sarcocystis cruzi antigens.

Sarcocystis neurona, a coccidian parasite shed by opossums (Didelphis spp.) in the Americas, is the major cause of equine protozoal myeloencephalitis (EPM) and induces disease in other domestic and wild animal species, including domestic dogs. Sarcocystis cruzi, despite its low pathogenicity for cattle (intermediate hosts), is worldwide distributed and uses mostly dogs as definitive hosts. The aims of this study were to test serological reactivities of dog sera to S. neurona and S. cruzi antigens, and to investigate potential serological cross-reactivity to these parasites. Sera from 353 Brazilian dogs were obtained from rural areas in the municipality of Ilhéus, Bahia, and examined by immunofluorescent antibody tests (IFAT). Antigens used in serological reactions consisted of S. neurona merozoites from a North American strain (SN138), and bradyzoites of S. cruzi obtained from Brazilian bovine hearts, with parasite species identity confirmed by PCR and sequencing of the 18S gene of the rDNA. Seropositivity to S. neurona and to S. cruzi were detected in 3.39% (12/353) and 4.81% (17/353) of the dogs, respectively. Ten canine sera reacted solely to S. neurona and 15 serum samples reacted only to S. cruzi. Two serum samples were simultaneously positive for both parasites. Sera from 14 dogs that tested positive by IFAT (9 for S. neurona and 3 for S. cruzi) and from two dogs that were negative by IFAT for the two parasites, were examined by Western blot using S. neurona as antigen; these sera reacted to a great number of protein bands, including antigens on the 16 and 30 KDa positions, which encompass immunodominant antigens for S. neurona in horses. Western blot did not show any specific pattern for S. neurona infection/exposure using canine sera. Dogs act as definitive hosts for several Sarcocystis spp. that infect farm animals, including horses, sheep, goats, water buffaloes and pigs, and for this reason, should contain antibodies to a broad repertoire of Sarcocystis spp. antigens. In conclusion, low percentages of dogs from rural areas of Ilhéus, Bahia, were reactive to both S. neurona and S. cruzi antigens. It is possible that other Sarcocystis species, besides S. neurona and S. cruzi, might have contributed for the seropositivity observed in this study. IFAT was more specific than Western blot to differentiate canine serological reactions to S. neurona and S. cruzi antigens.

The Podovirus ϕ80-18 Targets the Pathogenic American Biotype 1B Strains of Yersinia enterocolitica.

We report here the complete genome sequence and characterization of Yersinia bacteriophage vB_YenP_ϕ80-18. ϕ80-18 was isolated in 1991 using a Y. enterocolitica serotype O:8 strain 8081 as a host from a sewage sample in Turku, Finland, and based on its morphological and genomic features is classified as a podovirus. The genome is 42 kb in size and has 325 bp direct terminal repeats characteristic for podoviruses. The genome contains 57 predicted genes, all encoded in the forward strand, of which 29 showed no similarity to any known genes. Phage particle proteome analysis identified altogether 24 phage particle-associated proteins (PPAPs) including those identified as structural proteins such as major capsid, scaffolding and tail component proteins. In addition, also the DNA helicase, DNA ligase, DNA polymerase, 5′-exonuclease, and the lytic glycosylase proteins were identified as PPAPs, suggesting that they might be injected together with the phage genome into the host cell to facilitate the take-over of the host metabolism. The phage-encoded RNA-polymerase and DNA-primase were not among the PPAPs. Promoter search predicted the presence of four phage and eleven host RNA polymerase –specific promoters in the genome, suggesting that early transcription of the phage is host RNA-polymerase dependent and that the phage RNA polymerase takes over later. The phage tolerates pH values between 2 and 12, and is stable at 50°C but is inactivated at 60°C. It grows slowly with a 50 min latent period and has apparently a low burst size. Electron microscopy revealed that the phage has a head diameter of about 60 nm, and a short tail of 20 nm. Whole-genome phylogenetic analysis confirmed that ϕ80-18 belongs to the Autographivirinae subfamily of the Podoviridae family, that it is 93.2% identical to Yersinia phage fHe-Yen3-01. Host range analysis showed that ϕ80-18 can infect in addition to Y. enterocolitica serotype O:8 strains also strains of serotypes O:4, O:4,32, O:20 and O:21, the latter ones representing similar to Y. enterocolitica serotype O:8, the American pathogenic biotype 1B strains. In conclusion, the phage ϕ80-18 is a promising candidate for the biocontrol of the American biotype 1B Y. enterocolitica.

Isolation and molecular characterization of the first subgroup J avian leukosis virus from chicken in Pakistan

Since subgroup J avian leukosis virus (ALV-J) was first isolated in the United Kingdom in 1988, it has seriously hindered the development of the poultry industry worldwide. Although cases of ALV-J infection have been reported as early as 2001 in Pakistan, there was no further research on the isolation and molecular characteristics of ALVs. In the present study, we first isolated two ALVs from suspicious clinical samples that were collected from a desi chicken farm in Pakistan. The results of multiplex PCR and indirect immunofluorescent antibody assays confirmed that the two isolates (PK19FA01 and PK19SA01) belonged to ALV-J. The complete genomes of the two isolates were amplified, sequenced, and systematically analyzed. We found that gp85 of PK19FA01 was more similar to that of the prototype strain HPRS103, whereas gp85 of PK19SA01 was more similar to that of American strains. The two isolates contained an intact E element of 147 residues and had a unique 135 bp deletion in the redundant transmembrane of the 3′ untranslated region. The U3 region of the two isolates was highly homologous to that of American ALV-J strains. To our knowledge, this is the first report of the isolation, complete genome sequencing, and systematic molecular epidemiological investigation of ALV-J in Pakistan. Our findings could enrich epidemiological data and might contributed to more effective measures to prevent and control avian leukosis in Pakistan.

STAT3/SOCS3 axis contributes to the outcome of salmonid whirling disease.

There are differences in disease susceptibility to whirling disease (WD) among strains of rainbow trout. The North American strain Trout Lodge (TL) is highly susceptible, whereas the German Hofer (HO) strain is more resistant. The suppressor of cytokine signaling (SOCS) proteins are key in inhibiting cytokine signaling. Their role in modulating the immune response against whirling disease is not completely clear. This study aimed at investigating the transcriptional response of SOCS1 and SOCS3 genes to Myxobolus cerebralis along with that of several upstream regulators and immune response genes. M. cerebralis induced the expression of SOCS1, the IL-6-dependent SOCS3, the anti-inflammatory cytokine IL-10 and the Treg associated transcription factor FOXP3 in TL fish at multiple time points, which likely caused a restricted STAT1 and STAT3 activity affecting the Th17/Treg17 balance. The expression of SOCS1 and the IL-6-dependent SOCS3 was induced constraining the activation of STAT1 and STAT3 in TL fish, thereby causing Th17/Treg17 imbalance and leaving the fish unable to establish a protective immune response against M. cerebralis or control inflammatory reactions increasing susceptibility to WD. Conversely, in HO fish, the expression of SOCS1 and SOCS3 was restrained, whereas the expression of STAT1 and IL-23-mediated STAT3 was induced potentially enabling more controlled immune responses, accelerating parasite clearance and elevating resistance. The induced expression of STAT1 and IL-23-mediated STAT3 likely maintained a successful Th17/Treg17 balance and enabled fish to promote effective immune responses favouring resistance against WD. The results provide insights into the role of SOCS1 and SOCS3 in regulating the activation and magnitude of host immunity in rainbow trout, which may help us understand the mechanisms that underlie the variation in resistance to WD.

Transcriptome analysis of Drosophila melanogaster laboratory strains of different geographical origin after long-term laboratory maintenance.

Positive selection may be the main factor of the between‐population divergence in gene expression. Expression profiles of two Drosophila melanogaster laboratory strains of different geographical origin and long‐term laboratory maintenance were analyzed using microchip arrays encompassing probes for 18,500 transcripts. The Russian strain D18 and the North American strain Canton‐S were compared. A set of 223 known or putative genes demonstrated significant changes in expression levels between these strains. Differentially expressed genes (DEG) were enriched in response to DDT (p = .0014), proteolysis (p = 2.285E−5), transmembrane transport (p = 1.03E−4), carbohydrate metabolic process (p = .0317), protein homotetramerization (p = .0444), and antibacterial humoral response (p = 425E−4). The expression in subset of genes from different categories was verified by qRT‐PCR. Analysis of transcript abundance between Canton‐S and D18 strains allowed to select several genes to estimate their participation in latitude adaptation. Expression of selected genes was analyzed in five D. melanogaster lines of different geographic origins by qRT‐PCR, and we found two candidate genes that may be associated with latitude adaptation in adult flies—smp‐30 and Cda9. Quite possible that several alleles of these genes may be important for insect survival in the environments of global warming. It is interesting that the number of genes involved in local adaptation demonstrates expression level appropriate to their geographical origin even after decades of laboratory maintenance.

Characterization of a Borrelia miyamotoi membrane antigen (BmaA) for serodiagnosis of Borrelia miyamotoi disease

Borrelia miyamotoi is a tick-borne pathogen that causes Borrelia miyamotoi disease (BMD), an emerging infectious disease of increasing public health significance. B. miyamotoi is transmitted by the same tick vector (Ixodes spp.) as B. burgdorferi sensu lato (s.l.), the causative agent of Lyme disease, therefore laboratory assays to differentiate BMD from Lyme disease are needed to avoid misdiagnoses and for disease confirmation. We previously performed a global immunoproteomic analysis of the murine host antibody response against B. miyamotoi infection to discover antigens that could serologically distinguish the two infections. An initial assessment identified a putative lipoprotein antigen, here termed BmaA, as a promising candidate to augment current research-based serological assays. In this study, we show that BmaA is an outer surface-associated protein by its susceptibility to protease digestion. Synthesis of BmaA in culture was independent of temperature at either 23 °C or 34 °C. The BmaA gene is present in two identical loci harbored on separate plasmids in North American strains LB-2001 and CT13-2396. bmaA-like sequences are present in other B. miyamotoi strains and relapsing fever borrelia as multicopy genes and as paralogous or orthologous gene families. IgM and IgG antibodies in pooled serum from BMD patients reacted with native BmaA fractionated by 2-dimensional gel electrophoresis and identified by mass spectrometry. IgG against recombinant BmaA was detected in 4 of 5 BMD patient serum samples as compared with 1 of 23 serum samples collected from patients with various stages of Lyme disease. Human anti-B. turicatae serum did not seroreact with recombinant BmaA suggesting a role as a species-specific diagnostic antigen. These results demonstrated that BmaA elicits a human host antibody response during B. miyamotoi infection but not in a tested group of B. burgdorferi-infected Lyme disease patients, thereby providing a potentially useful addition for developing BMD serodiagnostic tests.