Sperm storage by females across the animal phyla: A survey on the occurrence and biomolecules involved in sperm storage.

Long-term sperm storage by females in various regions of the oviduct is documented across many invertebrate and vertebrate species. Although, many reports emphasize on the histology, histochemistry and ultrastructural features of sperm storage, very little is known about the mechanisms underlying the sperm storage. The current review documents the occurrence of sperm storage by females in a wide array of invertebrate and vertebrate species. This review also provides an insight on the presence of various molecular factors of the sperm storage tubules presumably responsible for the prolonged sperm storage with an emphasis on a model reptile, the Indian garden lizard, Calotes versicolor which contains a unique approximately 55-kDa protein in its utero-vaginal lavage and found to inhibit washed epididymal sperm motility in a concentration and time-dependent manner in a reversible fashion.

Constraining the Neutron Star Compactness: Extraction of the

The ^{23}Al(p,γ)^{24}Si reaction is among the most important reactions driving the energy generation in type-I x-ray bursts. However, the present reaction-rate uncertainty limits constraints on neutron star properties that can be achieved with burst model-observation comparisons. Here, we present a novel technique for constraining this important reaction by combining the GRETINA array with the neutron detector LENDA coupled to the S800 spectrograph at the National Superconducting Cyclotron Laboratory. The ^{23}Al(d,n) reaction was used to populate the astrophysically important states in ^{24}Si. This enables a measurement in complete kinematics for extracting all relevant inputs necessary to calculate the reaction rate. For the first time, a predicted close-lying doublet of a 2_{2}^{+} and (4_{1}^{+},0_{2}^{+}) state in ^{24}Si was disentangled, finally resolving conflicting results from two previous measurements. Moreover, it was possible to extract spectroscopic factors using GRETINA and LENDA simultaneously. This new technique may be used to constrain other important reaction rates for various astrophysical scenarios.

HD-Zip Proteins GL2 and HDG11 Have Redundant Functions in Arabidopsis Trichomes, and GL2 Activates a Positive Feedback Loop via MYB23.

The class IV homeodomain leucine zipper transcription factor GLABRA2 (GL2) acts in a complex regulatory circuit that regulates the differentiation of trichomes in Arabidopsis thaliana. We describe a genetic interaction with HOMEODOMAIN GLABROUS11 (HDG11), previously identified as a negative regulator of trichome branching. gl2 hdg11 double mutants display enhanced trichome cell-type differentiation defects. Transgenic expression of HDG11 using the GL2 promoter partially suppresses gl2 trichome phenotypes. Vice versa, expression of GL2 under the control of its native promoter partially complements hdg11 ectopic branching. Since gl2 hdg11 and gl2 myb23 double mutants and the triple mutant display similar trichome differentiation defects, we investigated a connection to the R2R3-MYB transcription factor MYB23. We show that MYB23 transcript levels are significantly reduced in shoots from gl2 mutants and that GL2 can drive the expression of a MYB23-promoter fusion to green fluorescent protein. Yeast one-hybrid, chromatin immunoprecipitation, and in planta reporter gene experiments indicate that an L1-box in the MYB23 promoter acts as a GL2 binding site. Taken together, our findings reveal a functional redundancy between GL2 and HDG11, two homeodomain leucine zipper transcription factors previously thought to mediate opposing functions in trichome morphogenesis. A model is proposed in which GL2 transcript levels are maintained through a positive feedback loop involving GL2 activation of MYB23.

Urogenital schistosomiasis among school children in northwestern Tanzania: Prevalence, intensity of infection, associated factors, and pattern of urinary tract morbidities.

Background: The planning and implementation of intervention measures against schistosomiasis, particularly mass administration, require knowledge of the current status of the infection. This is important for monitoring the impact of the intervention on disease indicators such as a decline in infection prevalence, intensity of infection, and urogenital morbidities. Following repeated rounds of mass treatment in northwestern Tanzania, the epidemiology of urogenital schistosomiasis has changed; thus, for the effective planning and allocation of resources, it is important to understand the current status of the disease in the targeted groups. Therefore, the objective of the current study was to determine the prevalence, intensity, and associated factors of Schistosoma haematobium infection and urinary tract morbidities in school-aged children from northwestern Tanzania. Materials and methods: An analytical cross-sectional study was conducted among schoolchildren aged 5-17 years between November and December 2022. A single urine sample was collected from each child and examined for the presence of S. haematobium eggs and microhaematuria using a urine filtration technique and a urine reagent dipstick. Each child underwent an ultrasonographic examination of the urinary tract according to the World Health Organization standards (Niamey protocol) to detect S. haematobium-related morbidities. Results: Of the 3225 participants, 54.2 % were female, and the mean age was 10.9 (±1.89) years. The overall prevalence of S. haematobium was 17.7 % (95 % CI: 16.4-19.1, 572/3225). Of the 572 infected children, 81.8 % (95 % CI: 78.4-84.9, 468/572) had light-intensity infections, and 18.2 % (95 % CI: 14.9-21.4, 104/572) had heavy-intensity infections. The prevalence of macro- and microhaematuria was 2.4 % (95 % CI: 1.9-3) and 18.5 % (95 % CI: 17.2-19.8), respectively. Age (aOR: 1.2, 95 % CI: 1.0-1.5), district of residence (aOR: 2.1, 95 % CI: 1.7-2.7) and history of schistosomiasis (aOR: 2.5, 95 % CI: 1.9-3.2) were significantly associated with urinary schistosomiasis infection. However, swallowing praziquantel during the last mass drug administration was protective (aOR 0.6, 95 % CI: 0.4-0.8). The overall prevalence of ultrasound-detectable urinary tract abnormalities was 9.9 % (95 % CI: 8.9-11.1, 299/2994) and included urinary bladder abnormalities in 9.9 % (95 % CI: 8.8-11, 297/2994), ureter abnormalities in 0.2 % (95 % CI: 0.07-0.4, 6/2994), and kidney abnormalities in 0.2 % (95 % CI: 0.09-0.4, 7/2994). Calcification of the urinary bladder was observed in 0.9 % (95 % CI: 0.6-1.3, 29/2994) of the examined children. Conclusions: Schistosoma haematobium infection is still prevalent among schoolchildren in the study setting, and it causes substantial morbidity at an early age. Transmission is driven by the age of the child, district of residence, and history of schistosomiasis. However, swallowing praziquantel in rounds of mass drug administration reduces transmission. Urogenital schistosomiasis infection is associated with haematuria and ultrasound-detectable morbidities. In S. haematobium endemic areas, routine ultrasound screening for urinary tract morbidities could be considered in annual mass treatment programmes for early management. Special attention should be given to children with proteinuria, microhaematuria, and heavy infection intensities.

Interferon release does not add discriminatory value to smear-negative HIV-tuberculosis algorithms.

Clinical algorithms for evaluating HIV-infected individuals for tuberculosis (TB) prior to isoniazid preventive therapy (IPT) perform poorly, and interferon-γ release assays (IGRAs) have moderate accuracy for active TB. It is unclear whether, when used as adjunct tests, IGRAs add any clinical discriminatory value for active TB diagnosis in the pre-IPT assessment. 779 sputum smear-negative HIV-infected persons, established on or about to commence combined antiretroviral therapy (ART), were screened for TB prior to IPT. Stepwise multivariable logistic regression was used to develop clinical prediction models. The discriminatory ability was assessed by receiver operator characteristic area under the curve (AUC). QuantiFERON-TB Gold in-tube (QFT-GIT) was evaluated. The prevalence of smear-negative TB by culture was 6.4% (95% CI 4.9-8.4%). Used alone, QFT-GIT and the tuberculin skin test (TST) had comparable performance; the post-test probability of disease based on single negative tests was 3-4%. In a multivariable model, the QFT-GIT test did not improve the ability of a clinical algorithm, which included not taking ART, weight <60 kg, no prior history of TB, any one positive TB symptom/sign (cough ≥ 2 weeks) and CD4+ count <250 cells per mm(3), to discriminate smear-negative culture-positive and -negative TB (72% to 74%; AUC comparison p=0.33). The TST marginally improved the discriminatory ability of the clinical model (to 77%, AUC comparison p=0.04). QFT-GIT does not improve the discriminatory ability of current TB screening clinical algorithms used to evaluate HIV-infected individuals for TB ahead of preventive therapy. Evaluation of new TB diagnostics for clinical relevance should follow a multivariable process that goes beyond test accuracy.

A mouse model of Leishmania braziliensis braziliensis infection produced by coinjection with sand fly saliva.

To development a reliable murine model of Leishmania braziliensis braziliensis infection, parasites were injected into BALB/c mice in the presence of phlebotomine sand fly salivary gland lysates, which have previously been shown to greatly increase the infectivity of L. major in mice. When injected with salivary gland lysates, L. braziliensis braziliensis produced progressively enlarging cutaneous nodules, containing many macrophages filled with Leishmania amastigotes. In contrast, L. braziliensis injected without gland extracts produced small and rapidly regressing lesions. Isoenzyme analysis, monoclonal antibodies, and the polymerase chain reaction with L. braziliensis-specific oligonucleotide primers and probes confirmed that parasites causing the lesions were L. braziliensis.

T cell and non-T cell compartments can independently determine resistance to Leishmania major.

In experimental murine cutaneous leishmaniasis caused by Leishmania major (Lm), the cellular determinants governing development of protective or exacerbative T cells are not well understood. We, therefore, attempted to determine the influence of T cell and non-T cell compartments on disease outcome. To this end, T cell chimeric mice were constructed using adult thymectomized lethally irradiated, bone marrow-reconstituted (ATXBM) animals of genetically resistant, C57BL/6, or susceptible, BALB/c, backgrounds. These hosts were engrafted with naive T cell populations from H-2-congenic susceptible, BALB.B6-H-2b, or resistant, C57BL/6.C-H-2d, animals, respectively. Chimeric mice were then infected with Lm, and disease outcome was monitored. BALB/c T cell chimeric mice, BALB/c ATXBM hosts given naive C57BL/6.C-H-2d T cells, resolved their infections as indicated by reductions in both lesion size and parasite numbers. Furthermore, the mice developed typical Th1 (interferon[IFN]-gamma hiinterleukin[IL]-4lo) cytokine patterns. In contrast, both sham chimeric, BALB/c ATXBM hosts given naive BALB/c T cells, and control irradiated euthymic mice succumbed to infection, producing Th2 profiles (IFN-gamma loIL-4hiIL-10hi). C57BL/6 T cell chimeras, C57BL/6 ATXBM hosts given naive BALB.B6-H-2b T cells, resolved their infections as did C57BL/6 sham chimeras and euthymic controls. Interestingly, whereas C57BL/6 control animals produced Th1 cytokines, chimeric animals progressed from Th0 (IFN-gamma hiIL-4hiIL-10hi) to Th2 (IFN-gamma loIL-4hiIL-10hi) cytokine profiles as cure ensued. Both reconstitution and chimeric status of all mice were confirmed by flow cytometry. In addition, T cell receptor V beta usage of Lm-specific blasts was determined. In all cases, V beta use was multiclonal, involving primarily V beta 2, 4, 6, 8.1, 8.2, 8.3, 10, and 14, with relative V beta frequencies differing between H-2b and H-2d animals. Most importantly, however, these differences did not segregate between cure and noncure outcomes. These findings indicate that: (a) genetic traits determining cure in Lm infection can direct disease outcome from both T cell and non-T cell compartments; (b) the presence of the curing genotype in only one compartment is sufficient to confer cure; (c) curing genotype T cells autonomously assume a Th1 cytokine profile-mediating cure; (d) noncuring genotype T cells can mediate cure in a curing environment, despite the onset of Th2 cytokine production; and lastly, (e) antigen specificity of responding T cells, as assessed by V beta T cell receptor diversity, is not a critical determinant of disease outcome.

Leishmania major-specific, CD4+, major histocompatibility complex class II-restricted T cells derived in vitro from lymphoid tissues of naive mice.

Several studies indicate that the outcome of experimental murine cutaneous leishmaniasis caused by Leishmania major (Lm) is determined by immunological events occurring shortly after infection. These events lead to outgrowth of either protective CD4+ T cells in the C57BL/6 mouse, which cures, or exacerbative cells in the BALB/c mouse, which succumbs to disease. Potential factors influencing the outgrowth of protective or exacerbative T cells include antigen-presenting cells (APC), cytokines, and parasite antigens. An in vitro system, in which one could precisely control the factors shaping early events in the T cell response to Lm, would be very useful. To this end, we have examined the in vitro response of naive lymphocytes to Lm promastigotes. The data presented here show that Lm-specific CD4+ T cell receptor alpha/beta + T cells can be generated in vitro from spleen and lymph node cell populations of naive mice. Furthermore, they can be obtained from the CD44low (unprimed) population of T lymphocytes, indicating that in vitro priming occurs. The ability to generate these T cells is dependent on the presence of live parasites and is not due to a parasite-derived nonspecific T cell mitogen. Restimulation, as assayed by proliferation, requires APC bearing syngeneic I-A. Optimal restimulation of the in vitro derived T cells is achieved only when live promastigotes are used. The T cells do not proliferate in response to a frozen-and-thawed lysate of promastigotes, yet they exhibit mild reactivity to lysates prepared from heat-shocked promastigotes. Furthermore, they do not recognize two predominant antigens on the promastigote surface, lipophosphoglycan and gp63. T cells derived in vitro with Lm show crossreactivity with live L. donovani, less crossreactivity with live L. mexicana, and no crossreactivity with live Bacillus-Calmette-Guerin or live Brugia malayi microfilariae. Finally, these early T cells, whether derived from healing C57BL/6 or nonhealing BALB/c mice, produce interleukin 2 (IL-2), IL-4, and interferon gamma.

Tumor necrosis factor plays a protective role in experimental murine cutaneous leishmaniasis.

The ability of mice to resist infection with L. major correlated directly with the capacity of their LNC to produce TNF in response to in vitro parasite challenge. Blocking TNF in vivo by passively administering anti-TNF antibodies exacerbated the course of L. major infection, resulting in substantially larger cutaneous lesions and elevated numbers of parasites within those lesions. In addition, treatment of infected mice with exogenous rHuTNF afforded host protection as evidenced by smaller lesion size and decreased parasite counts. Taken together, these results suggest a central role for TNF in resistance to L. major.

Saliva of the Lyme disease vector, Ixodes dammini, blocks cell activation by a nonprostaglandin E2-dependent mechanism.

Tick-borne pathogens would appear to be vulnerable to vertebrate host immune responses during the protracted duration of feeding required by their vectors. However, tick salivary components deposited during feeding may inhibit hemostasis and induce immunosuppression. The mode of action and the nature of immunosuppressive salivary components remains poorly described. We determined that saliva from the main vector of the agent of Lyme disease, Ixodes dammini, profoundly inhibited splenic T cell proliferation in response to stimulation with concanavalin A or phytohemagglutin, in a dose-dependent manner. In addition, interleukin 2 secretion by the T cells was markedly diminished by saliva. Tick saliva also profoundly suppressed nitric oxide production by macrophages stimulated with lipopolysaccharide. Finally, we analyzed the molecular basis for the immunosuppressive effects of saliva and discovered that the molecule in saliva responsible for our observations was not PGE2, as hypothesized by others, but rather, was a protein of 5,000 mol wt or higher.

Salivary gland lysates from the sand fly Lutzomyia longipalpis enhance Leishmania infectivity.

Leishmaniasis is a parasitic disease transmitted by phlebotomine sand flies. The role of sand fly saliva in transmission of the disease was investigated by injecting mice with Leishmania major parasites in the presence of homogenized salivary glands from Lutzomyia longipalpis. This procedure resulted in cutaneous lesions of Leishmania major that were routinely five to ten times as large and contained as much as 5000 times as many parasites as controls. With inocula consisting of low numbers of Leishmania major, parasites were detected at the site of injection only when the inoculum also contained salivary gland material. This enhancing effect of sand fly salivary glands on cutaneous leishmaniasis occurred with as little as 10 percent of the contents of one salivary gland of one fly. Material obtained from other bloodsucking arthropods could not mediate the phenomenon.

Regulation of differentiation to the infective stage of the protozoan parasite Leishmania major by tetrahydrobiopterin.

A critical step in the infectious cycle of Leishmania is the differentiation of parasites within the sand fly vector to the highly infective metacyclic promastigote stage. Here, we establish tetrahydrobiopterin (H4B) levels as an important factor controlling the extent of metacyclogenesis. H4B levels decline substantially during normal development, and genetic or nutritional manipulations showed that low H4B caused elevated metacyclogenesis. Mutants lacking pteridine reductase 1 (PTR1) had low levels of H4B, remained infectious to mice, and induced larger cutaneous lesions (hypervirulence). Thus, the control of pteridine metabolism has relevance to the mechanism of Leishmania differentiation and the limitation of virulence during evolution.

A method for the isolation and analysis of leucocytic cells from Leishmanial ear lesions in mice.

The standard model of experimental cutaneous leishmaniasis involves infection of mice with Leishmania major in a single footpad or the rump, and analysis of the subsequent immune response in draining lymph nodes. Relatively few studies have examined the lesion directly. Here, we describe a method for the isolation of cells from established leishmanial lesions in mouse ears. After physical disruption of lesion tissue and isolation of cells on density gradients, a variety of leucocytic cell phenotypes were identified by flow cytometry and cytology. The phenotypes of the viable cells obtained were similar, in proportion, to those observed in histologic sections of ear lesions. This technique may be useful for studying lesion-specific cell function within the first weeks after infection with Leishmania parasites.

A simple and effective method to assess murine delayed type hypersensitivity to proteins.

Mice sensitized with various protein antigens emulsified in complete Freund's adjuvant by a subcutaneous injection at the base of the tail exhibited both immediate and delayed type hypersensitivities 7-8 days following sensitization. Hypersensitivity was assessed by the degree of footpad swelling elicited by challenge with a heat-aggregated form of the antigen. The swelling response was specific for the antigen used for sensitization and often resulted in a near doubling of footpad thickness. Histological examination is conjunction with studies having passive transfer of serum and adoptive transfer of either T cell-enriched or T cell-depleted preparations indicated that whereas the immediate swelling response was achieved with serum, the delayed swelling response was mediated by T lymphocytes. Adoptive transfer of specific delayed type hypersensitivity was also accomplished with small numbers of lymph node T cells obtained following in vitro enrichment and propagation.

Reversion to virulence in Leishmania major correlates with expression of surface lipophosphoglycan.

An attenuated clone of Leishmania major was produced by chemical mutagenesis with N-methyl-N'-nitro-N-nitrosoguanidine and was biochemically characterized to determine the reason(s) for its loss of virulence. We found that the degree of virulence of L. major did not correlate with either the level of expression of promastigote surface protease (PSP) or with the enzymatic activity of the molecule. In contrast, the levels of lipophosphoglycan (LPG) expressed by the attenuated clone were found to be at least 6-fold less than those of virulent L. major. When the attenuated L. major was injected into BALB/c mice and allowed to revert to virulence, the degree of reversion to virulence that the parasites underwent correlated directly with the amount and form (metacyclic) of LPG expressed by the parasites. Thus, these results further implicate LPG as an important Leishmania virulence factor.

Aporphines as antagonists of dopamine D-1 receptors.

The aporphine alkaloids are a class of compounds known to possess activity at both D-1 and D-2 dopamine receptors. (R)-Apomorphine and (S)-bulbocapnine are examples of compounds which have agonist and antagonist activity, respectively, at D-1 receptors. A series of optically pure aporphines was synthesized and their activity at D-1 and D-2 dopamine receptors was studied. The (R)-aporphines uniformly had greater affinity for both D-1 and D-2 receptors than their S antipodes. Dihydroxy compound (R)-apomorphine, in accord with previous studies, was found to be a D-1 agonist. Aporphines possessing a single hydroxy group at C-11 are antagonists at the D-1 receptor. The corresponding methoxy compounds are virtually inactive at dopamine receptors. The most potent compounds, (R)-11-hydroxyaporphine (R-14) and (R)-10-bromo-11-hydroxyaporphine (R-26), are more potent than bulbocapnine as D-1 antagonists but are not as selective. A model for binding of aporphines to the D-1 receptor was formulated in which binding interactions between the receptor and the basic nitrogen and the C-11 hydroxy group of the aporphine are required for high-affinity binding to the receptor. The absolute configuration at C-6a determines the orientation of the N-6 lone pair and binding is optimal for the 6aR series. The agonist or antagonist activity of an aporphine is determined by the presence or absence, respectively, of a hydroxy group at C-10. A hydrophobic binding site may be present and may account for the high antagonist activity of (S)-bulbocapnine.

Resolution and absolute configuration of an ergoline-related dopamine agonist, trans-4,4a,5,6,7,8,8a,9-Octahydro-5-propyl-1H(or 2H)-pyrazolo[3,4-g]quinoline.

The title compound (+/-)-5 (R = Pro) (LY141865) has been resolved into a (-) isomer and a (+) isomer as the D- and L-tartrate salts, respectively. Biological studies have shown that dopamine agonist activity is a property of only the (-) isomer. Crystallographic analysis has proven that the absolute configuration of the active (-) isomer is the same as that of the natural ergolines.

Histologic characterization of experimental cutaneous leishmaniasis in mice infected with Leishmania braziliensis in the presence or absence of sand fly vector salivary gland lysate.

Leishmania braziliensis is the causative agent of human cutaneous leishmaniasis in parts of the New World. In the murine model of infection, L. braziliensis does not produce severe or lasting cutaneous lesions in either BALB/c or C3H mice. However, when the parasites are injected into BALB/c mice with salivary gland lysate of the sand fly vector for the parasite, infection is significantly enhanced, as measured by lesion size, parasite burden, and the outcome of infection. Histologic examination of these cutaneous lesions showed that initially, nodular and diffuse dermal infiltrates of neutrophils, eosinophils, and histiocytes occurred in all mice. Over time, the saliva-free lesions progressed to small organized granulomas of epithelioid macrophages that contained few parasites, with eventual resolution of inflammation and mild dermal fibrosis. The saliva-associated lesions progressed to extensive, poorly organized accumulations of heavily parasitized epithelioid macrophages, with persistent neutrophils and eosinophils, and minimal fibroplasia. These results indicate that sand fly salivary gland lysate markedly modifies the inflammatory response to infection with L. braziliensis.

Radiation injuries after fluoroscopic procedures.

Fluoroscopically guided diagnostic and interventional procedures have become much more commonplace over the last decade. Current fluoroscopes are easily capable of producing dose rates in the range of 0.2 Gy (20 rads) per minute. The dose rate often changes dramatically with patient positioning and size. Most machines currently in use have no method to display approximate patient dose other than the rough surrogate of total fluoroscopy time. This does not include patient dose incurred during fluorography (serial imaging or cine runs), which can be considerably greater than dose during fluoroscopy. There have been over 100 cases of documented radiation skin and underlying tissue injury, a large portion of which resulted in dermal necrosis. The true number of injuries is undoubtedly much higher. The highest dose procedures are complex interventions such as those involving percutaneous angioplasties, stent placements, embolizations, and TIPS. In some cases skin doses have been in excess of 60 Gy (6000 rads). In many instances the procedures have been performed by physicians with little training in radiation effects, little appreciation of the radiation injuries that are possible or the strategies that could have been used to reduce both patient and staff doses. Almost all of the severe injuries that have occurred were avoidable.

Environmental semivolatiles and nonvolatiles analysis by hyphenated spectral techniques.

Environmental sample extracts contain a variety of volatile and nonvolatile organic compounds exhibiting a range of polarities and concentrations. Although gas chromatography/mass spectrometry (GC/MS) is the method of choice thus far for such analyses, this technique used alone cannot adequately characterize the volatiles in such samples and is not amenable to environmental nonvolatiles. A more complete characterization of environmental and hazardous waste samples is required to assess the dangers posed to the nation's groundwater by hazardous waste dumps. Online spectral confirmation by directly linked GC/Fourier transform infrared (FTIR)/MS is shown to provide useful structural information on environmental volatiles in hazardous wastes, even when the analyte's spectrum is not in either spectral database. This information can lead to biological-hazard estimation. The diffuse reflectance Fourier transform infrared (DRIFT) technique, used in conjunction with thermospray MS or fast atom bombardment (FAB) MS, provides confirmed identifications or confirmed compound class assignments of organic nonvolatiles in solid wastes. This is believed to be the first report of spectral confirmation (identification or functionality) of organic volatiles and nonvolatiles in environmental samples.