A parasite reveals cryptic phylogeographic history of its host

This study compares the continental phylogeographic patterns of two wild European species linked by a host-parasite relationship: the field mouse Apodemus sylvaticus and one of its specific parasites, the nematode Heligmosomoides polygyrus. A total of 740 base pairs (bp) of the mitochondrial cytochrome b (cyt b) gene were sequenced in 122 specimens of H. polygyrus and compared with 94 cyt b gene sequences (974 bp) previously acquired for A. sylvaticus. The results reveal partial spatial and temporal congruences in the differentiation of both species' lineages: the parasite and its host present three similar genetic and geographical lineages, i.e. Western European, Italian and Sicilian, and both species recolonized northwestern Europe from the Iberian refuge at the end of the Pleistocene. However, H. polygyrus presents three particular differentiation events. The relative rate of molecular evolution of the cyt b gene was estimated to be 1.5-fold higher in the parasite than in its host. Therefore, the use of H. polygyrus as a biological magnifying glass is discussed as this parasite may highlight previously undetected historical events of its host. The results show how incorporating phylogeographic information of an obligate associate can help to better understand the phylogeographic pattern of its host.     

Pregnancy-associated murine protein-1 (PAMP-1) in wild rodents.

The presence of pregnancy-associated murine protein-1 (PAMP-1) in a number of wild rodent species was examined using tandem crossed immunoelectrophoresis and anti PAMP-1 antisera raised in rabbits. PAMP-1 is a protein present in female laboratory mice and rats; analogues were readily detected in the following species: Musculus domesticus, M. musculus, Apodemus sylvaticus, A. agrarius, A. flavicollis, and Microtus arvalus. Immunologically, cross-reacting proteins could not be detected in the laboratory hamster and laboratory guinea pig. Maternal PAMP-1 serum levels were recorded throughout pregnancy in the two species of house mouse: M. domesticus and M. musculus. Both proteins exhibited maximum concentration at day 11 of gestation. During early pregnancy, M. domesticus females had significantly higher serum levels than those of M. musculus females.     

Biochemical, physiological and morphological variation in unarmed hymenolepids (Eucestoda: Cyclophyllidae)

The genus Hymenolepis contains a number of unarmed species. These frequently possess similar morphologies and are difficult to discriminate using the traditional method of comparative morphology. A parasite of the long-tailed field mouse, Apodemus sylvaticus in northeast Ireland, resembles the widespread H. diminuta which is usually a parasite of the rat. Analysis of general and specific proteins of the adults in A. sylvaticus , laboratory mice and rats suggests that the parasite found in the former host and H. diminuta are genetically distinct, though more closely allied than either is to H. nana, H. citelli and H. microstoma . Experimental analysis of the growth and expulsion of the Irish material and H. diminuta from SPF C57 laboratory mice, rats and wild caught A. sylvaticus suggests that there are behavioural and physiological differences in these taxa. Both are expelled from C57 mice though the hymenolepid from Irish A. sylvaticus persists for 3 days more than those of H. diminuta . The former prospers better in rats than H. diminuta in A. sylvaticus . Detailed comparison of the gross morphology of cysticercoid and adult H. diminuta and the Irish hymenolepid reveals differences in size rather than qualitative attributes. The occurrence of H. diminuta in A. sylvaticus is discussed. It is concluded that the hymenolepid recovered from Irish A. sylvaticus differs sufficiently from H. diminuta to warrant species status and that it has adapted to the alimentary canal of A. sylvaticus . This cestode material is described under the name of H. hibernia sp. nov.     

Eimeviu (Protozoa: Coccidia) in wild populations of some British rodents

Four species of wild rodents from the British Isles, Myocustor coypus (the Coypu), Apodemus sylvaticus (the Wood mouse). Mus musculus (the House mouse) and Clethrionomys glureolus (the Bank vole) were examined during 1918 and 1919 for the presence of coccidian parasites of the genus Eimeriu. From an examination of 852 faecal samples a total of 14 species of Eimeriu was found. Species identification was based mainly on the morphology of the sporulated oocyst. The number of host species examined and the overall prevalence of infection were as follows: 252 M. coypus (45·6% infected), 471 A. sylvaticus 56·5%), 59 M. musculus (42·4%) and 70 C. glureolus (22·9%). The results are compared with previous surveys and discussed in the light of the general knowledge concerning coccidial infections. The present work showed that prevalence of infection in the wild rodents studied may be influenced by the age, weight and sex of the host, and by season.     

Phylogeography of a nematode (Heligmosomoides polygyrus) in the western Palearctic region: persistence of northern cryptic populations during ice ages?

This study establishes the continental phylogeographical pattern of a European nematode, Heligmosomoides polygyrus (Dujardin, 1845; Heligmosomoidea). We sequenced 687 base pairs of the mitochondrial DNA (mtDNA) cyt b gene for 136 individuals collected in 22 localities. The results revealed that H. polygyrus populations are separated into five major units corresponding to the Italian, northern European (Denmark and Ireland), Iberian, western European, and Balkan populations. Different subclades were also observed within the first two groups. Based on the rate of molecular evolution of H. polygyrus cyt b gene-estimated to 3.5%-3.7% divergence per million years (Myr) in a previous study--the isolation time of the five clades was estimated between 2.5 +/- 0.24 and 1.5 +/- 0.23 million years ago. Moreover, H. polygyrus presents a higher genetic variability in the Mediterranean peninsulas as compared to northwestern Europe, highlighting the role of these regions as refuge areas. Like its specific host, the wood mouse Apodemus sylvaticus, H. polygyrus' pattern of postglacial recolonization of northwestern Europe was initiated from Iberian populations, while Italian and Balkan populations did not expand to the north. The results also suggest the existence of forested and temperate refuges in the southern British Isles during the Quaternary. Finally, the genetic diversity as well as the level of genetic divergence between the lineages of H. polygyrus are compared to those observed in other vertebrate and invertebrate phylogeographical studies: the existence of highly differentiated lineages in H. polygyrus (5%-10% of genetic divergence) highlights that the effects of Pleistocene climate changes on free-living organisms are also reflected in their obligate parasites.     

On the interpretation of host-parasite ecology: Heligmosomoides polygyrus (Nematoda) in wild wood mouse (Apodemus sylvaticus) populations

This paper describes epidemiological and seasonal patterns in the interaction between wood mice, Apodemus sylvaticus and Heligmosomoides polygyrus. Data used in the analysis were collected by C. S. Elton and co-workers at Bagley Wood, Oxfordshire in the late 1920s. Heligmosomoides polygyrus was by far the most common helminth parasite with 70% of all wood mice infected and average intensity around 12 worms per mouse. Male and female mice were shown to harbour similar parasite burdens. Parasite numbers per host were highly overdispersed and were well described by the negative binomial distribution. There was little evidence for convexity in age (= weight)-intensity curves, either within or across sexes.
Host and parasite numbers showed predictable seasonal patterns, with mouse populations at their largest at the end of the breeding season, in August and September, and parasite populations at their largest in the late spring, around May. Results are discussed in relation to the ecology of H. polygyrus in wood and laboratory mice, and tentative comparison is made with human helminth infection. The interpretation of epidemiological patterns in these data was problematic. Of particular importance was the statistical distribution of parasites within the host population, and possible differences between mouse sexes in relation to growth, survival and trapping. Such difficulties are relevant to a range of similar field data.     

A genetic linkage map of the MSM Japanese wild mouse strain with restriction landmark genomic scanning (RLGS)

A high-resolution genetic map of the Mus musculus molossinus (MSM) Japanese wild mouse strain was constructed with restriction landmark genomic scanning (RLGS) and compared with that of the laboratory strain C3H. MSM is phylogenetically 1 million years apart from common laboratory mouse strains and is distinctly resistant to chemical carcinogenesis. Since it exhibits frequent genetic polymorphisms with laboratory mice but can still be easily crossed with laboratory strains, hybrids between MSM and carcinogen-sensitive laboratory mouse strains provide excellent materials for analysis of modifier genes and genetic changes during carcinogenesis. We have generated MSM backcross progeny with the C3H strain, which is extremely sensitive to hepatocarcinogenesis, to construct the present map. RLGS profiles with two combinations of restriction enzymes (NotI–PvuII–PstI, NotI–PstI–PvuII) yielded more than 2000 spots each. The polymorphism rate was about 39.2%, and of a total of 1732 polymorphic spot loci identified, 1371 could be assigned to specific chromosomes by comparison with 79 microsatellite marker loci. Thus, 1450 loci, on all chromosomes except for Y, effectively mapped 90% of the genome (1431.7 cM length). Although some spots might be derived from the same NotI site, each NotI site potentially generating two fragments, the presence of at least 515 loci groups with different progeny distribution patterns dispersed through the genome with an average spacing of 3 cM, means that this genetic map should be useful for analysis of various biological phenomena, including carcinogenesis and ontogenesis, at the gene level. Received: 25 August 1999 / Accepted: 20 December 1999     

Intraspecies polymorphism of Cryptosporidium parvum revealed by PCR-restriction fragment length polymorphism (RFLP) and RFLP-single-strand conformational polymorphism analyses

A glycoprotein (Cpgp40/15)-encoding gene of Cryptosporidium parvum was analyzed to reveal intraspecies polymorphism within C. parvum isolates. Forty-one isolates were collected from different geographical origins (Japan, Italy, and Nepal) and hosts (humans, calves, and a goat). These isolates were characterized by means of DNA sequencing, PCR-restriction fragment length polymorphism (PCR-RFLP), and RFLP-single-strand conformational polymorphism (RFLP-SSCP) analyses of the gene for Cpgp40/15. The sequence analysis indicated that there was DNA polymorphism between genotype I and II, as well as within genotype I, isolates. The DNA and amino acid sequence identities between genotypes I and II differed, depending on the isolates, ranging from 73.3 to 82.9% and 62.4 to 80.1%, respectively. Those among genotype I isolates differed, depending on the isolates, ranging from 69.0 to 85.4% and 54.8 to 79.2%, respectively. Because of the high resolution generated by PCR-RFLP and RFLP-SSCP, the isolates of genotype I could be subtyped as genotypes Ia1, Ia2, Ib, and Ie. The isolates of genotype II could be subtyped as genotypes IIa, IIb, and IIc. The isolates from calves, a goat, and one Japanese human were identified as genotype II. Within genotype II, the isolates from Japan were identified as genotype IIa, those from calves in Italy were identified as genotype IIb, and the goat isolate was identified as genotype IIc. All of the genotype I isolates were from humans. The Japanese isolate (code no. HJ3) and all of the Nepalese isolates were identified as genotypes Ia1 and Ia2, respectively. The Italian isolates were identified as genotype Ib, and the Japanese isolate (code no. HJ2) was identified as genotype Ie. Thus, the PCR-RFLP-SSCP analysis of this glycoprotein Cpgp40/15 gene generated a high resolution that has not been achieved by previous methods of genotypic differentiation of C. parvum.     

Molecular and biological diversity of HIV-1 in Brazil.

To determine the genomic polymorphism and biological properties present in HIV-1 Brazilian isolates, we analyzed five viral isolates obtained from patients residing in Rio de Janeiro (P1 and P5), S?o Paulo (P3) and Bahia (P2 and P4) states. For each viral isolate in vitro characteristics such as replication rate, syncytium-inducing capacity and cell death were observed in lymphoblastoid (H9, CEM and peripheral blood mononuclear cells) as well as monocytoid (U937) cells. In addition, the evaluation of the restriction fragment length polymorphism of these isolates was also performed using a panel of endonucleases such as Hind III, Bgl II, Sac I, Pst I, Kpn I and Eco RI. One of the isolates (P1), showed the highest phenotypic and genotypic divergence, when compared to others. The results found suggest a HIV heterogeneity in Brazil similar to that already described in other regions of the world.     

小家鼠和实验小鼠遗传特性的比较研究

本文用同工酶电泳法、微量细胞毒法和免疫双向扩散法对我国4个动物地理区的6个采集点的156个小家鼠(Mus musculus)进行了遗传特性的调查。结果发现:在全部被测的13个位点中,小家鼠在7个位点上存在着多种实验小鼠中罕见的基因组成;而不同动物地理区和亚区的小家鼠的遗传特性又各不相同。从而指出将小家鼠的特有基因导入实验小鼠,培育新品系的重大意义。     

Restriction enzyme banding of mouse metaphase chromosomes

Fixed metaphase chromosomes from mouse strain RIII embryos or A9 cells were treated with a restriction endonuclease, followed by Giemsa staining. Aha I, Hinf I, or Mbo I treatment produced a C-band pattern, and Eco RII or Hae III produced a G-band plus C-band pattern. Ava II and Bst NI each produced a G-band pattern, but on most chromosomes only a small segment of each C-band, adjacent to the centromere, was stained. These tiny residual C-bands may contain a minor satellite located adjacent to the major satellite clusters.     

Intraspecies Polymorphism of Cryptosporidium parvum Revealed by PCR-Restriction Fragment Length Polymorphism (RFLP) and RFLP-Single-Strand Conformational Polymorphism Analyses

A glycoprotein (Cpgp40/15)-encoding gene of Cryptosporidium parvum was analyzed to reveal intraspecies polymorphism within C. parvum isolates. Forty-one isolates were collected from different geographical origins (Japan, Italy, and Nepal) and hosts (humans, calves, and a goat). These isolates were characterized by means of DNA sequencing, PCR-restriction fragment length polymorphism (PCR-RFLP), and RFLP-single-strand conformational polymorphism (RFLP-SSCP) analyses of the gene for Cpgp40/15. The sequence analysis indicated that there was DNA polymorphism between genotype I and II, as well as within genotype I, isolates. The DNA and amino acid sequence identities between genotypes I and II differed, depending on the isolates, ranging from 73.3 to 82.9% and 62.4 to 80.1%, respectively. Those among genotype I isolates differed, depending on the isolates, ranging from 69.0 to 85.4% and 54.8 to 79.2%, respectively. Because of the high resolution generated by PCR-RFLP and RFLP-SSCP, the isolates of genotype I could be subtyped as genotypes Ia1, Ia2, Ib, and Ie. The isolates of genotype II could be subtyped as genotypes IIa, IIb, and IIc. The isolates from calves, a goat, and one Japanese human were identified as genotype II. Within genotype II, the isolates from Japan were identified as genotype IIa, those from calves in Italy were identified as genotype IIb, and the goat isolate was identified as genotype IIc. All of the genotype I isolates were from humans. The Japanese isolate (code no. HJ3) and all of the Nepalese isolates were identified as genotypes Ia1 and Ia2, respectively. The Italian isolates were identified as genotype Ib, and the Japanese isolate (code no. HJ2) was identified as genotype Ie. Thus, the PCR-RFLP-SSCP analysis of this glycoprotein Cpgp40/15 gene generated a high resolution that has not been achieved by previous methods of genotypic differentiation of C. parvum.     

Two distinct endogenous type C viruses isolated from the asian rodent Mus cervicolor: conservation of virogene sequences in related rodent species.

The cocultivation of a lung cell line from the Southeast Asian mouse Mus cervicolor with cells from heterologous species has resulted in the isolation of two new distinct type C viruses. Both viruses are endogenous to M. cervicolor and are present in multiple copies in the cellular DNA of these mice. One of the viruses, designated M. cervicolor type CI, replicates readily in the SIRC rabbit cell line and is antigenically related to the infectious primate type C viruses isolated from a woolly monkey (simian sarcoma-associated virus) and gibbon apes (gibbon ape leukemia virus). This virus is also closely related by both immunological and nucleic acid hybridization criteria to a type C virus previously isolated from a second Asian murine species, Mus caroli. The isolation of the M. cervicolor type C I virus thus provides further evidence that the infectious primate type C viruses originated by trans-species infection of primates by an endogenous virus of mice. The second virus, designated M. cervicolor type C II, replicates well in various cell lines derived from the laboratory mouse Mus musculus. While antigenically related to type C viruses derived from M. musculus, the M. cervicolor type C II virus isolate can be readily distinguished from standard murine leukemia viruses. Both new type C viruses from M. cervicolor are unrelated to the previously described retrovirus (M432) isolated from the same Mus species. The DNA of M. cervicolor therefore contains multiple copies of at least three distinct classes of endogenous viral genes. An examination of the cellular DNA of other rodent species for nucleic acid sequences related to the genomes of both M. cervicolor type C I and II reveals that both viruses have been highly conserved evolutionarily, and that other species of rodents, such as laboratory mice and rats, contain endogenous virogenes related to those in the DNA of M. cervicolor.     

Naturally occurring murine leukemia viruses in wild mice: characterization of a new "amphotropic" class.

A new class of murine leukemia viruses, isolated from wild Mus musculus trapped in California, is described. These viruses, designated "amphotropic," replicate in mouse, rabbit, mink, human, guinea pig, and rat cells, but not in hamster, quail, or duck cells. They show N-tropism for mouse cells, and do not trigger the XC cell response. They are distinct by interference and virus neutralization testing from the previously recognized mouse-tropic and xenotropic MuLV classes. Mouse-tropic viruses occuring along with three of the four amphotropic isolates were found to be distinguishable by virus neutralization from other mouse-tropic murine leukemia virus strains of laboratory mouse origin.     

Haemoglobin variants in mice.

Haemoglobin variants were studied in wild and laboratory house mice (Mus musculus), including standard and new inbred strains, using starch-gel electrophoretic technique. Single (Hbbs) or diffuse (Hbbd) types of haemoglobin were found in all of them. The embryonic haemoglobin pattern was different from although similar to that of the adult in all the strains. The haemoglobins revealed monomorphism in the inbred strains, while polymorphism was observed in non-inbred laboratory and wild mice.     

The Original Pink-Eyed Dilution Mutation (P) Arose in Asiatic Mice: Implications for the H4 Minor Histocompatibility Antigen, Myod1 Regulation and the Origin of Inbred Strains

Allelic variation of the mouse pink-eyed dilution (p) gene in common laboratory strains and wild mice was examined by Southern blot and by polymerase chain reaction. In these assays the original p mutation allele found in strains SJL/J, 129/J, B10.129(21m), P/J and FS/Ei most closely matches an Asian Mus musculus allele, confirming anecdotal accounts of the Asian origin of this mutation. In contrast, the wild-type allele found in other common laboratory strains was apparently derived from Mus domesticus. Analysis of chromosome 7 loci both proximal and distal to the p locus demonstrates that strains SJL/J, 129/J, B10.129(21M), P/J and FS/Ei contain DNA segments of varying length derived from M. musculus. Strains 129/J and B10.129(21M) contain the largest segment of M. musculus-derived DNA (about 5 cM), including the loci Myod1, p, three clustered GABA(A) receptor subunit loci (Gabrg3, Gabra5 and Gabrb3), and Snrpn. The difference in the species origin of genes from this region of chromosome 7 may underlie the basis of the antigenicity of the minor histocompatibility antigen H4, defined by the strain B10.129(21M), and may account for the enhanced Myod1 activity observed in SJL/J mice.     

Mammary tumorigenesis in feral mice: identification of a new int locus in mouse mammary tumor virus (Czech II)-induced mammary tumors.

A population of Mus musculus subsp. musculus (Czech II), recently isolated from the wild, lack endogenous mouse mammary tumor virus (MMTV) proviral genomes. Some of these mice carry an infectious MMTV [designated MMTV (Czech II)] that is transmitted in the milk and is associated with mammary tumor development. This virus is distinct from laboratory strains of MMTV present in inbred mice. An MMTV (Czech II) genome was found within a 0.5-kilobase region of the cellular genome in five of 16 Czech II mammary tumors. MMTV insertion at this site activates expression of a 2.4-kilobase species of RNA from a previously silent cellular gene. This region of the cellular genome was designated int-3 since it is unrelated to the int-1 and int-2 loci. The int-3 locus does not appear to correspond to other proto-oncogenes but is well conserved among mammalian species.     

Identification of enterohepatic Helicobacter species by restriction fragment-length polymorphism analysis of the 16S rRNA gene

Background. Restriction fragment-length polymorphism (RFLP) analysis of a 1,200-bp polymerase chain reaction–amplified DNA fragment of gene coding for 16S rRNA was used to generate restriction profiles of 11 enterohepatic Helicobacter spp. isolated from various animals and humans.
Methods. The amplicon from each Helicobacter sp. was digested with four restriction endonucleases: Alu I, Hinf I, Hha I, and Dde I. Alu I digestion produced five patterns that were useful for initial differentiation.
Results. Most Helicobacter spp. isolated from rodents had the same RFLP profiles by Alu I digestion (except H. rodentium and H. cholecystus ), but they had different RFLP profiles by Hha I digestion. Only H. bilis and " H. rappini" mouse isolates could not be readily distinguished by the polymerase chain reaction-RFLP method. However, these two species can be distinguished using H. bilis specific primers. Some of the Helicobacter spp. have an intervening sequence in their 16S rRNA gene, which changes the RFLP patterns; in these cases, sequencing is the preferred method to make an appropriate diagnosis.
Conclusions. The RFLP method used in this study was straightforward and rapid and should prove useful as an adjunct for identification and classification of multiple enterohepatic Helicobacter spp.     

Development of unique house mouse resources suitable for evolutionary studies of speciation

Two house mouse subspecies, Mus musculus domesticus and Mus musculus musculus, form a hybrid zone in Europe and represent a suitable model for inferring the genes contributing to isolation barriers between parental taxa. Despite long-term intensive studies of this hybrid zone, we still know relatively little about the causes and mechanisms maintaining the 2 taxa as separate subspecies; therefore, to gain insight into this process, we developed 8 wild-derived inbred house mouse strains. In order to produce strains as pure domesticus or musculus genomes as possible, the individuals used to establish the breeding colony for the 3 domesticus and 2 of the musculus strains were captured in the Czech Republic from wild populations at extreme western and eastern edges of the subspecific contact zone, respectively. The remaining 3 musculus strains were bred from mice captured about 250 km east of the hybrid zone. Genetic analysis based on 361 microsatellite loci showed that 82% of these markers are diagnostic for either the musculus or the domesticus strains. In order to demonstrate the potential utility of this genetic differentiation in such strains, phenotypic variation was scored for 2 strains from opposite edges of the hybrid zone and significant differences in morphology, reproductive performance, in vitro immune responses, mate choice based on urinary signals, and aggressiveness were found. In addition, the 3 strains derived from musculus populations far from the hybrid zone display significant differences in polymorphism in hybrid male sterility when crossed with the laboratory strains C57BL/6 or C57BL/10, which have a predominantly domesticus genome. Although further studies will be necessary to demonstrate intersubspecific differences, all analyses presented here indicate that these newly developed house mouse strains represent a powerful tool for elucidating the genetic basis of isolation barriers in hybrid zones and for studying speciation in general.