Chronic hepatitis B (CHB) is caused by the Hepatitis B virus (HBV) and affects millions of people worldwide. Developing an effective CHB therapy requires using in vivo screening methods, such as mouse models reflecting CHB based on hydrodynamic delivery of plasmid vectors containing a replication-competent HBV genome. However, long-term expression of HBV proteins is accompanied by production of progeny virions, thereby requiring a Biosafety Level (BSL) 3 animal facility. In the present study, we introduced a point mutation in the START codon of the HBV polymerase to develop a mouse model reflecting chronic hepatitis B infection without formation of viral progeny. We induced the mouse model by hydrodynamic injection of adeno-associated virus plasmid vector (pAAV) and minicircle plasmid (pMC) constructs into C57Bl/6 and C3H/HeN mouse strains, monitoring HBV antigens and antibodies in blood by enzyme-linked immunosorbent assay and analyzing liver expression of HBV core antigen by immunohistology. Persisting expression of viral antigens over 140 days (study endpoint) was observed only in the C3H/HeN mouse strain when using pAAV/1.2HBV-A and pMC/1.0HBV-D with pre-C and pre-S recombination sites. In addition, pAAV/1.2HBV-A in C3H/HeN sustained HBV core antigen positivity up to the study endpoint in C3H/HeN mice. Moreover, introducing the point mutation in the START codon of polymerase effectively prevented the formation of viral progeny. Our study establishes an accessible and affordable experimental paradigm for developing a robust mouse model reflecting CHB suitable for preclinical testing of anti-HBV therapeutics in a BSL2 animal facility.
Borrelia burgdorferi sensu lato (s.l.) is the etiological agent of Lyme disease, transmitted by ticks of the genus Ixodes Latreille. Diagnosis of Lyme disease in humans is often difficult and a detailed knowledge of the circulation of B. burgdorferi s.l. in tick hosts is therefore fundamental to support clinical procedures. Here we developed a molecular approach for the detection of B. burgdorferi s.l. in North Italian Ixodes ricinus (Linnaeus). The method is based on the amplification of a fragment of the groEL gene, which encodes a heat-shock protein highly conserved among B. burgdorferi s.l. species. The tool was applied in both qualitative and Real-time PCR approaches testing ticks collected in a North Italian area. The obtained results suggest that this new molecular tool could represent a sensitive and specific method for epidemiological studies aimed at defining the distribution of B. burgdorferi s.l. in I. ricinus and, consequently, the exposure risk for humans.
Monoclonal antibody specific for an epitope of cretory-secretory antigen protein of Opisthorchis felineus (Rivolta, 1884) (Trematoda: Opisthorchiidae) with a molecular weight of 28 kDa was used in a sandwich enzyme-linked immunosorbent assay (ELISA) for immobilisation of liver fluke specific antigen to the solid phase. Examination of human sera by this ELISA compared with commercial assays demonstrated that the monoclonal antibody epitope is located within this significant parasite protein. Anti-idiotypic antibody specific for the paratope of this monoclonal antibody was obtained by a hybridoma technique. Mimicking an epitope of excretory-secretory antigen of O. felineus, it had the capacity to bind specific antibody and elicit an antibody response. The value of anti-idiotypic antibody as a substitute for the liver fluke antigen was tested by ELISA using serum samples of infected dogs. Anti-idiotypic antibody proved to be of value in both an indirect-ELISA and a competitive-ELISA for diagnosis of opisthorchiasis. Mature trematodes were isolated from all infected animals. The faecal egg counts were negative in dogs with a relatively small number of parasites, despite finding antibodies in serum by ELISA. Substitution of parasite antigen with anti-idiotype avoids the use of experimental animals and also reduces time-consuming steps of antigen preparation., Aitbay K. Bulashev, Sergey N. Borovikov, Shynar S. Serikova, Zhanbolat A. Suranshiev, Vladimir S. Kiyan, Saule Z. Eskendirova., and Obsahuje bibliografii
Lepeophtheirus simplex Ho, Gómez et Fajer-Avila, 2001 is a parasite of Sphoeroides annulatus (Jenyns), an economically important fish species, with potential for aquaculture, in northwestern Mexico. The goal of this study was to describe the developmental stages under experimental conditions and seasonal fecundity of this parasite on wild fish. There are two naupliar, one copepodid, two chalimus and two pre-adult stages preceding the adult of L. simplex. The results support previous findings, which point out that the life cycle of the caligid copepods includes only six post-naupliar stages. The generation time from egg extrusion to adult for L. simplex was approximately 10 days at 22 °C. The body length of the ovigerous females ranged between 2.2 and 4.1 mm, and its fecundity between 12 and 36 eggs per string. Fecundity was negatively correlated with the egg size and positively correlated with the egg string length. Our data did not reveal significant differences in fecundity among sampling months, but ovigerous females were significantly larger in March (when water temperature was 22 °C) than in June and July (when water temperature was 30 °C). To some extent, our fecundity results contrast with those found in species of sea lice from higher latitudes. Undoubtedly, biological information on different species of sea lice from different environmental conditions will enhance our understanding of their infection strategies and will be valuable, given the increasing interest in marine fish farming in Mexico., Francisco Neptalí Morales-Serna, Ana Inés Rivas-Salas, Samuel Gómez, Emma Josefina Fajer-Ávila., and Obsahuje bibliografii
The Natal multimammate mouse, Mastomys natalensis, occurs throughout sub-Saharan Africa. Mitochondrial phylogenetics indicate this species was fragmented during the Pleistocene, forming six matrilineage phylogroups: A-I, A-II, A-III, B-IV, B-V, B-VI with distinct ranges. All except the A-III lineage are identified as natural reservoirs of mammarenaviruses. M. natalensis A-III is found in western Ethiopia and is the only lineage reported in the country. While screening 203 small mammal samples from Dhati Welel National Park for mammarenaviruses, we detected mammarenavirus RNA in nine samples, eight from M. natalensis and one from M. awashensis. A sequence similarity search and phylogenetic analysis confirmed the M. natalensis mitochondrial DNA belongs to the A-III lineage. We characterised the complete virus genome, which showed typical mammarenavirus organisation. Phylogenetic analysis indicated it clusters with Gairo virus found in M. natalensis B-IV in Tanzania, while showing sufficient divergence from other mammarenaviruses to be considered as a new species, for which we proposed the name Dhati Welel. Additional sampling in the M. natalensis A-III phylogeographic range should help determine whether the detection of the virus in M. awashensis represents a local spill-over or if the virus circulates in both Mastomys species.
Congenital toxoplasmosis is reportable disease in Europe. To prevent it antibody serological tests were introduced in several European countries as a part of screening programmes. Immunoglobulin G (IgG) avidity index testing is one of these tests for diagnosing acute infection with Toxoplasma gondii (Nicolle et Manceaux, 1908) in pregnant women. However, a low or moderate IgG avidity index can give inconclusive results for predicting woman's status. From June 2012 until the end of 2014, 17,990 women were included in the national screening program to prevent congenital toxoplasmosis. One hundred and twenty-six women were consecutively included in the study because they had low or moderate IgG avidity. Every woman with possible acute toxoplasmosis was followed up every month till delivery. Fifty-eight of 126 (46%) women got infected in months before current pregnancy, 39 women (31%) were infected early in pregnancy. Twenty-nine pregnant women of 126 (23%) got infected in the second/third trimester of pregnancy. New cut off for IgG avidity index was 0.11. With this cut off, we were able to exclude T. gondii acute infection in the first trimester with very good diagnostic accuracy (area under the curve (AUC) = 0.95, 95% confidence Interval (CI) 0.91-0.99, sensitivity 0.95, specificity 0.86). If an IgG avidity index above 0.11 is measured in a woman's serum and she is in the first trimester of pregnancy, then a odds ratio (OR) for acute infection with T. gondii is below 1 (OR 0.11, 95% CI 0.05-0.25, P < 0.0001). If we measure IgG avidity index that is ≥ 0.11 in the first trimester of pregnancy, we can exclude infection with T. gondii with good diagnostic accuracy in our cohort of women. With a new cut off we could reduce number of invasive procedures such as amniocentesis and put less pregnant women in distress.
Diclofenac is a drug commonly used in human and veterinary medicine for the treatment of diseases associated with inflammation and pain. Medicinal products enter waste and surface waters on an everyday basis and contaminate the aquatic environment. Fish are therefore permanently exposed to these chemicals dissolved in their aquatic environment. To simulate variable environmental conditions, the aim of our study was to examine adverse effects of diclofenac under different temperatures of cell incubation (18, 21, 24, 27 and 30 °C). Cytotoxic and -static effects of diclofenac in concentrations of 0.001 μg/ml, 0.01 μg/ml, 0.1 μg/ml, 1 μg/ml, 10 μg/ml and 100 μg/ml for the carp (Cyprinus carpio) cultured leukocytes were quantified using detection of lactate dehydrogenase released from damaged cells. Overall DCF cytotoxicity was relatively low and its impact was pronounced at higher temperature and DCF concentration. Cells growth inhibition is changing more rapidly but it is high mainly at the highest concentration from low temperature. DNA fragmentation was not detected in tested leukocyte cell line. CYP450 increased diclofenac cytotoxicity only at the highest concentration but at incubation temperatures 18 and 27 °C. Leukocyte viability is essential for immune functions and any change can lead to reduction of resistance against pathogens, mainly in cold year seasons, when the immune system is naturally suppressed.
The diet composition and prey selection of grey wolves (Canis lupus) inhabiting the Roztocze and Solska Forest (south-east Poland) was studied based on an analysis of scats collected in 2001-2002 (n = 84) and 2017-2020 (n = 302). In both periods, wolves preyed mainly on wild ungulates (96.5-96.7% of consumed biomass). Roe deer (Capreolus capreolus) was the most critical wolf prey accounting for 57.8% of consumed biomass in 2001-2002 and 49.2% and 2017-2020, but wolves positively select only wild boar (Jacob's selectivity index D = 0.213 in 2001-2002 and 0.710 in 2017-2020) and fallow deer (D = 0.588 only in 2017-2020). The largest species – moose Alces alces and red deer Cervus elaphus – were consumed less than expected from their share in the ungulate community. Predation on medium-sized wild mammals and domestic animals was low, 0.8-2.2% and 1.1-2.7% of the biomass consumed, respectively. The breadth of the wolf diet was very narrow and identical in both study periods (B = 1.07), while the similarity of diet composition was high (α = 0.999). This study indicated the stability of the wolf diet over two decades and the importance of wild boar as a food source for this carnivore.
Some moth larvae feed not on plants but on keratin and/or chitin produced by animals. These substances are polymers and are commonly found in bird nests as feathers and raptor pellets. Many qualitative studies have examined the association of keratin/chitin feeding moths with bird nests. However, few studies have quantified the species composition with respect to type of nest and habitat. Hence, we have studied the degree to which the niches of these moths differ in terms of type of nest and habitat. We set-up open-top nest boxes for the Ural owl Strix uralensis (damp exposed nests from which owl chicks were fledged successfully) and artificial bird nests (mesh bags filled with duck down to imitate dry exposed nests) in a deciduous forest and artificial bird nests in an urban setting in Aichi Prefecture, central Japan, and collected the contents of the nests every two months from June to December 2014. We recorded 592 individuals of five keratin/chitin feeding moth species (Tineidae, Tineinae) from the contents. Using non-metric multidimensional scaling and cluster analysis of the relative abundances of individual species in each type of nest in forest and urban settings, these species were classified into three groups: (1) Monopis longella and M. congestella inhabiting forest, (2) M. flavidorsalis and Niditinea tugurialis mainly in damp exposed nests in forest and (3) N. piercella mainly in dry exposed nests in urban areas. This classification was compared with findings of other studies. As a result, these moths probably have different niches with respect to nest type (damp or dry), keratin/chitin as a source of food (raptor pellets or feathers), and habitat (forest or urban area). Furthermore, we suggest that the evolution of larviparity in M. congestella might be related to its preference for feathers as a source of food for the development of its larvae.
Management of foreign-currency household debt in Romania in the aftermath of the Global Financial Crisis in 2008 had the effect of deepening pre-2008 class disparities and treated debtor categories differently according to their income. In this article we contribute to the debate on subaltern financialisation by showing how post-crisis credit and housing policies contributed to the fact that today different debtor groups (i.e. by type of credit but also by time of lending) find themselves at opposing ends of the political spectrum based on different class alliances, with those who benefited from the crisis-management polices positioning themselves against those who were the ‘losers’ under these same policies.