Recombinant congenic strains derived from A/J and C57BL/6J: a tool for genetic dissection of complex traits

Complex genetic traits can be dissected in mice, using well-defined sets of recombinant inbred strains, congenic strains, and recombinant congenic strains (RCS). We report the creation of a series of 37 independent RCS derived from the commonly used inbred strains of laboratory mouse A/J (A) and C57BL/6J (B6). These RCS were derived by systematic inbreeding of independent pairs of animals from a (F1 x A) x A and a (F1 x B) x B double backcross (N3), to create AcB and BcA strains, respectively. Fifteen AcB strains and 22 BcA strains at between 18 and 30 generations of inbreeding have been generated, are healthy, and show stable breeding performance. These strains have been genotyped for a total of 625 informative microsatellite DNA markers covering the entire genome, with an average spacing of 2.6 cM. Haplotype analyses indicate that on average, AcB and BcA strains contain 13.25% of the donor genome, a value close to the 12.5% expected from the breeding scheme used in their creation. In the AcB set, approximately 79% of the B6 genome has been transferred in independent strains, while in the BcA set approximately 84% of the A genome is represented on the B6 background. This represents an excellent coverage of congenic segments from both parental genomes in the two sets of strains, which can now be used to map simple and complex traits in a genome-wide fashion. As an example of the power of AcB/BcA strains as a mapping tool, the 37 strains were typed for susceptibility to infection with Legionella pneumophila, a monogenic trait controlled by the Lgn1 locus on Chromosome 13. Analysis of the strain distribution pattern of L. pneumophila susceptibility allowed direct mapping of Lgn1 to a 3-cM interval. The AcB/BcA set should prove a useful tool with which to investigate the complex genetic basis of known interstrain differences between A and B6 for many important diseases.     

Cutting edge: susceptibility to the larval stage of the helminth parasite Taenia crassiceps is mediated by Th2 response induced via STAT6 signaling

Using STAT6(-/-) BALB/c mice, we analyzed the role of STAT6-induced Th2 response in determining the outcome of murine cysticercosis caused by the helminth parasite Taenia crassiceps. After T. crassiceps infection, wild-type BALB/c mice developed a strong Th2-like response; produced high levels of IgG1, IgE, IL-4, as well as IL-13; and remained susceptible to T. crassiceps. In contrast, similarly infected STAT6(-/-) mice mounted a strong Th1-like response; produced high levels of IgG2a, IL-12, IFN-gamma, as well as nitric oxide; and efficiently controlled T. crassiceps infection. These findings demonstrate that Th2-like response induced via STAT6-mediated signaling pathway mediates susceptibility to T. crassiceps and, furthermore, that unlike the case in most helminths, immunity against T. crassiceps is mediated by a Th1-like rather than Th2-like response.     

Different effects of chorionic gonadotropin on Taenia crassiceps and Taenia solium cysticerci cultured in vitro

Hormones play a significant role in murine Taenia crassiceps cysticercosis, and they may also participate in the susceptibility to Taenia solium cysticercosis. In the present study, in vitro effects are reported for human chorionic gonadotropin (hCG) on the larval stages of T. crassiceps (WFU strain) and T. solium. hCG effectively promotes parasite reproduction, i.e., it increases the number of buds on T. crassiceps cysticerci and the percentage of evagination and parasite length in T. solium. This is the first report in which a direct effect of hCG is reported for a parasite. hCG or mouse luteinizing hormone could be recognized by the cysticerci as mitogenic factors and contribute to the female and pregnancy bias toward susceptibility to T. crassiceps and T. solium cysticercosis, respectively.     

Genetic basis for the psychostimulant effects of nicotine: a quantitative trait locus analysis in AcB/BcA recombinant congenic mice

Genetic differences in sensitivity to nicotine have been reported in both animals and humans. The present study utilized a novel methodology to map genes involved in regulating both the psychostimulant and depressant effects of nicotine in the AcB/BcA recombinant congenic strains (RCS) of mice. Locomotor activity was measured in a computerized open-field apparatus following subcutaneous administration of saline (days 1 and 2) or nicotine on day 3. The phenotypic measures obtained from this experimental design included total basal locomotor activity, as well as total nicotine activity, nicotine difference scores, nicotine percent change and nicotine regression residual scores. The results indicated that the C57BL/6J (B6) were insensitive to nicotine over the entire dose-response curve (0.1, 0.2, 0.4 and 0.8 mg/kg). However, the 0.8-mg/kg dose of nicotine produced a significant decrease in the locomotor activity in the A/J strain and a wide and continuous range of both locomotor excitation and depression among the AcB/BcA RCS. Single-locus association analysis in the AcB RCS identified quantitative trait loci (QTL) for the psychostimulant effects of nicotine on chromosomes 11, 12, 13, 14 and 17 and one QTL for nicotine-induced depression on chromosome 11. In the BcA RCS, nicotine-induced locomotor activation was associated with seven putative regions on chromosomes 2, 7, 8, 13, 14, 16 and 17. There were no overlapping QTL and no genetic correlations between saline- and nicotine-related phenotypes in the AcB/BcA RCS. A number of putative candidate genes were in proximity to regions identified with nicotine sensitivity, including the alpha2 subunit of the nicotinic acetylcholine receptor and the dopamine D3 receptor.     

Preferential growth of Taenia crassiceps cysticerci in female mice holds across several laboratory mice strains and parasite lines

A retrospective study of our 14-yr records on experimental Taenia crassiceps (ORF(fast) line) cysticercosis (n = 1,198) shows that in 16 of 17 different mice strains, female mice are more frequently infected and carry larger individual parasite loads than males. However, sexual differences in parasite loads significantly varies between strains in relation to their different genetic backgrounds (BALB > C57Bl = OTHERS > C3H). The coefficient of variation in all female mice is significantly smaller than that of all males, an indication of males' more potent, but erratically effective, restraint of cysticercus growth. Similar positive growth bias for female mice is shown by other lines of cysticerci, i.e., HYG(slow) and WFU(slow). These results contravene the usual expectation of female hosts being more resistant than males to parasite infections, and they point to the multiple factors that combined determine sex related differences of mice to experimental cysticercosis infection.     

Genetic control of high density lipoprotein-cholesterol in AcB/BcA recombinant congenic strains of mice

Epidemiological studies show that high HDL-cholesterol (HDLc) decreases the risk of cardiovascular disease. To map genes controlling lipid metabolism, particularly HDLc levels, we screened the plasma lipids of 36 AcB/BcA RC mouse strains subjected to either a normal or a high-fat/cholesterol diet. Strains BcA68 and AcB65 showed deviant HDLc plasma levels compared with the parental A/J and C57BL/6J strains; they were thus selected to generate informative F2 crosses. Linkage analyses in the AcB65 strain identified a locus on chromosome 4 (Hdlq78) responsible for high post-high fat diet HDLc levels. This locus has been previously associated at genome-wide significance to two regions in the human genome. A second linkage analysis in strain BcA68 identified linkage in the vicinity of a gene cluster known to control HDLc levels. Sequence analysis of these candidates identified a de novo, loss-of-function mutation in the ApoA1 gene of BcA68 that prematurely truncates the ApoA1 protein. The possibility of dissecting the specific effects of this new ApoA1 deficiency in the context of isogenic controls makes the BcA68 mouse a valuable new tool.     

Early removal of alternatively activated macrophages leads to Taenia crassiceps cysticercosis clearance in vivo

To determine the role of alternatively activated macrophages in modulating the outcome of experimental cysticercosis caused by Taenia crassiceps, we investigated the effect of removal of alternatively activated macrophage by injecting clodronate-loaded liposomes into susceptible BALB/c mice. Following T. crassiceps infection, mice receiving PBS-loaded liposomes developed a dominant Th2-type response associated with the presence of alternatively activated macrophages together with antigen-specific hyporesponsiveness and high parasite burden. In contrast, similarly infected mice treated with clodronate-loaded liposomes mounted a mixed Th1/Th2-type response, reversed antigen-specific hyporesponsiveness and did not carry notable alternatively activated macrophage populations. These factors were associated with increased resistance to T. crassiceps cysticercosis. Interestingly, early AAM? depletion was enough to limit parasite growth. However, if macrophages were depleted late in the infection, no effect on parasite burden was observed. These findings demonstrate that alternatively activated macrophages play a critical role in mediating susceptibility to experimental cysticercosis in which their early recruitment may favor parasite survival.     

Genetic analysis of the psychostimulant effects of nicotine in chromosome substitution strains and F2 crosses derived from A/J and C57BL/6J progenitors

Previous research utilizing the AcB/BcA recombinant congenic strains (RCS) of mice mapped provisional quantitative trait loci (QTLs) for the psychostimulant effects of nicotine to multiple regions on chromosomes 7, 11, 12, 14, 16, and 17. The current study was designed to confirm these QTLs in an A/J (A) × C57Bl/6J (B6) F2 cross and a panel of B6.A chromosome substitution strains (CSS). The panel of B6.A CSS consists of 21 strains, each carrying a different A/J chromosome on a B6 background. The A × B6 F2, CSS, A, and B6 mice were tested for sensitivity to the effects of nicotine on locomotor activity using a computerized open-field apparatus. In A × B6 F2 mice two QTLs were identified which confirm those previously observed in the AcB/BcA RCS. Significant differences in the expression of nicotine-induced activity were associated with loci on chromosome 11 (D11Mit62) and chromosome 16 (D16Mit131) in the A × B6 F2. At the chromosome 11 QTL, an A allele was associated with lower nicotine-induced activity scores relative to the B6. In contrast, the A allele was associated with greater relative nicotine activity values for the chromosome 16 QTL. A survey of the CSS panel confirmed the presence of QTLs for nicotine activation on chromosomes 2, 14, 16, and 17 previously identified in the AcB/BcA RCS. In the informative CSS strains, A alleles were consistently associated with greater nicotine-induced activity scores compared to the B6. The results of the present study are the first to validate QTLs for sensitivity to the effects of nicotine across multiple strains of mice. QTLs on chromosomes 2, 11, 14, 16, and 17 were confirmed in CSS and/or F2 mice. Significantly, the identification of a QTL on chromosome 16 has now been replicated in three crosses derived from the A and B6 progenitors.     

Genetic control of susceptibility to carcinogen-induced colorectal cancer in mice: The Ccs3 and Ccs5 loci regulate different aspects of tumorigenesis

Colorectal cancer (CRC) is a multistep disease that involves a two-way interaction between a complex genetic pre-disposition component, and a set of poorly understood extrinsic environmental factors. In mice, CRC can be induced by treatment with azoxymethane (AOM). Using a set of AcB/BcA recombinant congenic strains derived from CRC-susceptible A/J and CRC-resistant C57Bl/6J (B6) progenitors, we previously detected the Ccs3 locus (colon cancer susceptibility locus 3) as a major regulator of CRC susceptibility. Phenotyping of additional AcB/BcA strains for susceptibility to AOM-induced CRC has refined the Ccs3 interval to a 6.7 Mb segment on chromosome 3. In addition, the presence of intermediate susceptibility phenotypes in individual AcB/BcA strains suggested additional gene effects regulating CRC susceptibility in A/J and B6 strains. Those were investigated by linkage analysis and whole genome scanning in a set of 208 informative (B6 x A/J)F2 progeny, using tumor multiplicity as a quantitative measure of susceptibility. This analysis validated the important role of Ccs3 in regulating this trait, and additionally detected contribution from a second locus on the distal portion of chromosome 9 (LOD = 3.76), that was given the temporary designation of Ccs5. Ccs5 modulates tumor multiplicity in F2 animals bearing at least one A/J-derived susceptibility allele at Ccs3, with A/J-derived Ccs5 susceptibility alleles being inherited in a recessive manner. There is a strong additive effect of Ccs3 and Ccs5 on tumor multiplicity in F2 mice: mice doubly homozygotes for A/J or B6 alleles at Ccs3 and Ccs5 show tumor numbers similar to those of parental A/J and B6, respectively. Interestingly, the Ccs5 region overlaps several quantitative trait loci previously reported to regulate intestinal homeostasis and susceptibility to intestinal colitis in mice and humans. Our findings identify a novel two-locus system regulating CRC susceptibility in mice, of which the relevance to human CRC can now be tested experimentally.     

Protection from murine cysticercosis by immunization with a parasite cysteine protease

Central nervous system infection by Taenia solium cysts causes neurocysticercosis, a common neurological infection in the Third World. We have previously isolated cysteine proteases from Taenia crassiceps and T. solium. In this study, we immunized BALB/c mice with the purified T. solium cysteine protease and challenged them with Taenia crassiceps. Immunized mice had a 72% reduction in parasite burden compared to mice that received no immunization. Immunized mice developed antigen specific lymphocyte proliferation. These data support further studies of the T. solium cysteine protease as a vaccine candidate.     

The growth of the metacestodes of Taenia crassiceps in white mice.

The effect of the sex of the intermediate host on the growth and development of the metacestodes of Taenia crassiceps has been investigated. Three different strains of mice were used, the C R S, L A C A and C F L P strains and two strains of metacestode, the Toi and the E R S strains. The results show that matacestodes of T. crassiceps grow and multiply more rapidly in female than in male mice, although both sexes are equally susceptible to the infection. The origin of the parasite, i.e., from a rat or mouse host, affects the parasite growth.     

Sex steroids and parasitism: Taenia crassiceps cisticercus metabolizes exogenous androstenedione to testosterone in vitro

Sex hormones are known to modulate immune responses and may be implicated in sex associated susceptibilities to infections. Taenia crassiceps cysticerci grow to larger numbers in female mice than in males. Gonadectomy alters the course of this infection and hormone replacement with 17beta-estradiol increases the parasite numbers. However, in chronic Taenia crassiceps cysticercosis the sex-hormone profile of males becomes more like that of the females' and progressively loose their sexual behavior. To have further insight in these outstanding endocrinological effects induced by the parasite upon the host, we investigated the parasite's capacity to produce sex steroids. In vitro experiments showed that Taenia crassiceps cysticerci transform 3H-Androstenedione to 3(H)-Testosterone, but not 3H-Pregnenolone. The production of 3H-Testosterone increased when the parasite numbers doubled. A recrystallisation procedure demonstrated that the metabolite identified by TLC was in fact testosterone. Thus, the cysticercus has the ability to use 3H-Androstenedione to make Testosterone possibly by a 17beta-Hydroxysteroid deshidrogenase-like activity in the parasite. In vivo, the parasite could use steroid precursors from the host to produce sex hormones, either accidentally or as needed for its own development, and thus alters the host's normal environment with sexual and immunological repercussions.     

Altered T helper responses in CD40 and interleukin-12 deficient mice reveal a critical role for Th1 responses in eliminating the helminth parasite Taenia crassiceps

A key feature of helminth infections is the induction of strong Th2-biased immune responses in their hosts. We have previously found that Th2-like responses mediate susceptibility to the helminth parasite Taenia crassiceps, probably by inhibiting Th1 responses required for the development of protective immunity against this parasite. Here we show that mice lacking interleukin-12p35 (IL-12p35-/-) following T. crassiceps infection, failed to mount a Th1 response, but developed a strong Th2-type response, produced higher levels of IgG1, IgE, interleukin-4, interleukin-5 as well as interleukin-13 than wild-type mice, and became highly susceptible to the larval stage of this cestode. In contrast, similarly-infected CD40 deficient BALB/c mice (CD40-/-) displayed impairment of both Th1 and Th2-type responses associated with low levels of interferon-gamma as well as IgE, interleukin-4, interleukin-5 and interleukin-13, but efficiently controlled T. crassiceps infection. Together, these findings suggest a detrimental role for Th2-biased responses during the larval stage of T. crassiceps infection. Furthermore, they also suggest a pivotal role for CD40 in developing Th2-type responses.     

Absence of the SP/SP receptor circuitry in the substance P-precursor knockout mice or SP receptor, neurokinin (NK)1 knockout mice leads to an inhibited cytokine response in granulomas associated with murine Taenia crassiceps infection

Neurocysticercosis, caused by the cestode Taenia solium, is the most common parasitic infection of the human central nervous system that leads to seizures. Taenia crassiceps cysticercosis in mice is an experimental model for Taenia solium cysticercosis. Similar to the human infection, live parasites cause little or no granulomatous inflammation. Dying parasites initiate a granulomatous reaction. The neuropeptide, substance P (SP), stimulates T-helper (TH) 1 cytokine production. In the current studies, we determined whether absence of SP/SP receptor circuitry in the SP-precursor, preprotachykinin, knockout or SP-receptor, neurokinin (NK) 1, knockout mice affected granuloma cytokine production. We hence compared the levels of Th1 cytokines interleukin (IL)-2 and interferon (IFN)-gamma, and levels of Th2/immunoregulatory cytokines IL-4 and IL-10, by enzyme-linked immunosorbent assay in T. crassiceps-induced granulomas derived from infected C57BL/6 wild type (WT) versus SP-precursor knockout and NK1 knockout mice. We found that mean levels of IL-2, IFN-gamma, IL-4, and IL-10 in infected WT-derived granulomas were significantly higher than those of granulomas derived from infected SP-precursor knockout or the NK1 receptor (NKIR)knockout mice. Levels of Th2/immunoregulatory cytokines, IL-4 and IL-10 were higher in early stage granulomas (histologically-staged on basis of evidence of parasite remnants) versus late stage granulomas (no parasite-remnants) of both knockouts, whereas the reverse was noted in WT-derived granulomas. These study established that the absence of an SP/SP receptor circuitry in the SP precursor knockout mice or NK1 receptor knockout mice led to an inhibited cytokine response.     

Genetic analysis of alcohol intake in recombinant inbred and congenic strains derived from A/J and C57BL/6J progenitors

The objective of the present study was to map quantitative trait loci (QTL) for alcohol intake using A × B/B × A recombinant inbred (RI) and AcB/BcA recombinant congenic (RC) strains of mice that were independently derived from the A/J and C57BL/6J progenitors. Mice were screened for levels of alcohol consumption with four days of forced exposure to alcohol, followed by three weeks of free choice between water and a 10% alcohol solution. Alcohol consumption data previously collected for 27 A × B/B × A RI strains were reanalyzed using a larger marker set and composite interval mapping. The reanalysis found markers on Chromosome 2 (D2Mit74, 107 cM) (males and females) and on Chromosome 11 (Pmv22, 8 cM) (females only) that exceeded the threshold for significant loci, and found suggestive loci (in males) on Chromosomes 10 (D10 Mit126, 21 cM), 12 (D12Mit37, 1 cM), 15 (Pdgfb, 46.8 cM), and 16 (D16Mit125, 29 cM). An additional suggestive locus was identified in female RI mice on Chromosome 11 (D11Mit120, 47.5 cM). Composite interval mapping (CIM) analysis indicated that there was a significant association between loci at Pdgfb and D2Mit74 in both males and females. Analysis of the AcB/BcA RC strains identified 11 QTL on Chromosomes 2, 3, 5,6, 7, 8, 9, 10, 12, 13, and 15. QTL on Chromosomes 7, 10, 12, and 15 were identified in both the A × B/B × A RI and AcB/BcA RC strains of mice. Additional QTLs identified on Chromosomes 2, 3, 7, 11, and 15 overlap with those previously identified in the literature using strains of mice with a C57BL/6J progenitor.     

Inhibitory role of antibodies in the development of Taenia solium and Taenia crassiceps toward reproductive and pathogenic stages.

Untreated Taenia solium cysticerci obtained from different naturally infected pigs vary notably in their capacity to develop into intestinal tapeworms in prednisolone-treated hamsters, whereas cells derived from Taenia crassiceps cysticerci after 2 mo of infection almost always develop to cysticerci in the peritoneal cavity of susceptible BALB/cAnN mice. Preincubation of whole cysticerci or parasite cells with mice immunoglobulins raised against an 18-mer peptide epitope (GK-1) common to both parasites significantly interferes with both transformations. These crippling effects of antiparasite antibodies suggest new forms of immunological interference with parasite biology other than simple killing. Antibodies that cripple biological functions of the parasite, e.g., their development to reproductive or pathogenic stages, make them important protagonists in taeniasis/cysticercosis disease as classic parasitocidal antibodies. Different serum levels of crippling antibodies in the infected pigs could be responsible for the varied ability of cysticerci to convert to tapeworms. Antigens capable of inducing crippling antibodies, e.g., GK-1, could be useful as a therapeutic vaccine for pigs in order to reduce parasite transmission.     

Infrapopulation dynamics of a wild strain of Taenia crassiceps (WFU) (Cestoda: Taeniidae) in BALB/cJ mice

Taenia crassiceps cysticerci form large infrapopulations that persist in the tissues of their rodent hosts. Early infrapopulation growth appears inhibited and is followed by rapid increases that appear not to be controlled by the host immune response. This investigation was undertaken to examine the infrapopulation growth dynamics of a normally developing strain (WFU) of T. crassiceps during a 60-day primary intraperitoneal (i.p.) infection. Three, 6, 9, 14, 28, and 60 days after i.p. inoculation of 5 cysticerci, mice were killed, and the numbers of larvae, developmental stage, and buds per larva were recorded. Larval infrapopulation abundance increased exponentially beginning on day 6 postinoculation (PI), indicating an initial lag in reproduction. A stage-structured exponential growth model, assuming no mortality, fits the larval infrapopulation dynamics in terms of the numbers of larvae in reproductive and nonreproductive stages, indicating that cysticerci evade or suppress (or both) host immune mechanisms that are parasite restrictive after the first week of infection.     

Protective immunity against Taenia crassiceps murine cysticercosis induced by DNA vaccination with a Taenia saginata tegument antigen

This study investigated the protective capacity of the recombinant Taenia saginata Tso18 antigen administered as a DNA vaccine in the Taenia crassiceps murine model of cysticercosis. This Tso18 DNA sequence, isolated from a T. saginata oncosphere cDNA library, has homologies with Taenia solium and Echinococcus sp. It was cloned in the pcDNA3.1 plasmid and injected once intramuscularly into mice. Compared to saline-vaccinated control mice, immunization reduced the parasite burden by 57.3-81.4%, while lower levels of non-specific protection were induced in control mice injected with the plasmid pcDNA3.1 (18.8-33.1%) or a plasmid with irrelevant construct, pcDNA3.1/3D15 (33.4-38.8%). Importantly, significant levels of protection were observed between the pcDNA3.1/Tso18 plasmid and pcDNA3.1/3D15 plasmid immunized mice. Mice immunized with pTso18 synthesized low levels of, primarily IgG1 sub-class, antibodies. These antibodies were shown to recognize a 66 kDa antigen fraction of T. crassiceps and T. solium. Splenocytes enriched in both CD4+CD8- and CD4-CD8+ T cells from these vaccinated mice proliferated in vitro when exposed to antigens from both T. solium and T. crassiceps cestodes. Immunolocalization studies revealed the Tso18 antigen in oncospheres of T. saginata and T. solium, in the adult tapeworm and in the tegument of T. solium cysticerci. The protective capacity of this antigen and its extensive distribution in different stages, species and genera of cestodes points to the potential of Tso18 antigen for the possible design of a vaccine against cestodes.     

Effect of oral zinc supplementation upon Taenia crassiceps murine cysticercosis.

The effect of zinc supplementation on Taenia crassiceps murine cysticercosis was studied in susceptible BALB/cAnN mice. Female offspring of mice supplemented with high zinc throughout gestation and lactation were intraperitoneally infected with T. crassiceps cysticerci. Offspring from nonsupplemented mothers were used as controls. Significantly fewer parasites were recovered from zinc-supplemented mice (Zsm) 30 days after infection. Increased resistance was not related to the IgG antibody response. At early stages of infection, T cells from Zsm proliferated to T. crassiceps antigens, whereas cells from control mice did not respond. Infection caused in both groups a decrease in CD3+ cell percentages, which was more pronounced in the controls, and paralleled by a decrease in CD8+ cells; CD3+ and CD8+ percentages returned to normal levels at later stages of infection. In contrast, the CD4+ subpopulation only decreased in control mice. Intracellular cytokine determinations indicate that zinc supplementation favored a stronger and persistent type-1 T cell response in cysticerci-infected mice, which probably participates in the observed increased resistance.     

Mapping of clinical and expression quantitative trait loci in a sex-dependent effect of host susceptibility to mouse-adapted influenza H3N2/HK/1/68

Seasonal influenza outbreaks and recurrent influenza pandemics present major challenges to public health. By studying immunological responses to influenza in different host species, it may be possible to discover common mechanisms of susceptibility in response to various influenza strains. This could lead to novel therapeutic targets with wide clinical application. Using a mouse-adapted strain of influenza (A/HK/1/68-MA20 [H3N2]), we produced a mouse model of severe influenza that reproduces the hallmark high viral load and overexpression of cytokines associated with susceptibility to severe influenza in humans. We mapped genetic determinants of the host response using a panel of 29 closely related mouse strains (AcB/BcA panel of recombinant congenic strains) created from influenza-susceptible A/J and influenza-resistant C57BL/6J (B6) mice. Combined clinical quantitative trait loci (QTL) and lung expression QTL mapping identified candidate genes for two sex-specific QTL on chromosomes 2 and 17. The former includes the previously described Hc gene, a deficit of which is associated with the susceptibility phenotype in females. The latter includes the phospholipase gene Pla2g7 and Tnfrsf21, a member of the TNFR superfamily. Confirmation of the gene underlying the chromosome 17 QTL may reveal new strategies for influenza treatment.