Toxoplasma gondii in lactating animals: potential risk to milk consuming population in Khyber Pakhtunkhwa.

Toxoplasma gondii is an intracellular zoonotic protozoan parasite usually infects human and animal worldwide. This study aimed to analyze the sero-prevalence of T. gondii in blood of lactating animals and human living in close proximity and also to detect Toxoplasma DNA in unpasteurized milk of the studied animals. A total of 233 blood and milk samples were collected from lactating animals, and 735 blood samples were taken from humans in District Upper Dir, Khyber Pakhtunkhwa, Pakistan. The blood samples were analyzed through ELISA while the milk samples were analyzed by PCR for the presence of T. gondii DNA. A standard questionnaire was introduced to collect the data from the participants. In animals, the reported sero-prevalence was 32.18% for IgM, 17.16% for IgG, and 6.4% for both IgM and IgG. The reported positivity for T. gondii DNA in milk was 14.44%, 34.8%, 20%, and 26% in sheep, goats, cows, and buffaloes, respectively. In the human blood samples, 9.8% were found positive for IgM and 11.2% for IgG while none of the samples was found positive for both IgM and IgG. Overall sero-prevalence reported in females was significantly higher than the male (p<0.05) poor hygiene condition (p < 0.0001) were the significant risk factors associated with T. gondii infections in animals. In conclusion, T. gondii infection is prevalent in lactating animals and humans using their raw milk in the study area. It is suggested that raw milk should be considered as a vehicle for the transmission of T. gondii to humans. Proper pasteurization of milk is very useful in limiting the transmission of infection. Awareness and control programs should be implemented to prevent the infection.

Sprouty 2 binds ESCRT-II factor Eap20 and facilitates HIV-1 gag release.

The four ESCRT (endocytic sorting complexes required for transport) complexes (ESCRT-0, -I, -II, and -III) normally operate sequentially in the trafficking of cellular cargo. HIV-1 Gag trafficking and release as virus-like particles (VLPs) require the participation of ESCRTs; however, its use of ESCRTs is selective and nonsequential. Specifically, Gag trafficking to release sites on the plasma membrane does not require ESCRT-0 or -II. It is known that a bypass of ESCRT-0 is achieved by the direct linkage of the ESCRT-I component, Tsg101, to the primary L domain motif (PTAP) in Gag and that bypass of ESCRT-II is achieved by the linkage of Gag to ESCRT-III through the adaptor protein Alix. However, the mechanism by which Gag suppresses the interaction of bound ESCRT-I with ESCRT-II is unknown. Here we show (i) that VLP release requires the steady-state level of Sprouty 2 (Spry2) in COS-1 cells, (ii) that Spry2 binds the ESCRT-II component Eap20, (iii) that binding Eap20 permits Spry2 to disrupt ESCRT-I interaction with ESCRT-II, and (iv) that coexpression of Gag with a Spry2 fragment that binds Eap20 increases VLP release. Spry2 also facilitated release of P7L-Gag (i.e., release in the absence of Tsg101 binding). In this case, rescue required the secondary L domain (YPX(n)L) in HIV-1 Gag that binds Alix and the region in Spry2 that binds Eap20. The results identify Spry2 as a novel cellular factor that facilitates release driven by the primary and secondary HIV-1 Gag L domains.

Rydberg energies and transition probabilities of Li I for np-ms (m ≤ 5) transitions.

Background: Mathematical modeling provides grounds for understanding scientific systems theoretically. It serves as a guide for experimentalists in determining directions of investigation. Recently, the Covid-19 pandemic has caused disturbances in almost every walk of life. Scientists have played their role and have continued research on the effects of the pandemic. Various mathematical models have been used in different branches of science (Djilali et al. in Phys Scr 96 12 124016, 2021; Math Biosci Eng 18(6):8245-8256, 2021; Zeb et al. in Alex Eng J 61(7):5649-5665). Well-established mathematical models give results close to those obtained by experiments. The Weakest Bound Electron Potential Model is one such model, which explains hydrogen-like atoms and ions. This model has been used extensively for hydrogen-like atoms and ions to calculate energies of Rydberg levels and ionization energies. This model has been used extensively for hydrogen-like atoms and ions to calculate energies of Rydberg levels and ionization energies. Results: This paper presents the energies of the Rydberg series, 2s2ns, and 2s2np of Li I, calculated using WBEPM. The energies are used to calculate transition probabilities from np to 2s, 3s, 4s, and 5s levels. The transition probabilities are compared with corresponding values in published data where available. The agreement with known values is good; most of the transition probabilities calculated in this work are new. A computer program was developed to find the value of the dipole matrix element. The calculations were further verified by calculating the lifetimes of some low-lying levels. Conclusions: Four series of Li I have been studied, and energies of the Rydberg levels in the series were calculated. The energies then are used to calculate transition probabilities from np to ms transitions, where m = 2, 3, 4, & 5 and n = 1-15. The results are compared where available. An excellent agreement with previously published data shows the reliability of calculations. Most of the transition probabilities are new.

Human-black bear conflict: crop raiding by Asiatic black bear (Ursus thibetanus) in Azad Jammu and Kashmir, Pakistan.

Asiatic black bear has long been in conflict with human beings crop raiding is a major cause of this conflict frequently noted in South Asia. Crops raided by black bears affected by temporal, spatial and anthropogenic attributes. Insight in this conflict and its mitigation is vital for the conservation of this threatened species. Present study aimed to evaluate crop raiding by black bears in the mountainous region of Azad Jammu and Kashmir. Field surveys were carried out to observe spatial and temporal crop raiding features between 2015-2020 and data gathered using designed questionnaires randomly tailored in villages nearby the forests. Results revealed that maize was the sole crop raided by black bears. A total of 28-acre area was raided by black bear in the fall season (Aug-November) resulting in a damage of 51 metric tons, whole raiding was carried out at night. Each respondent received crop damage on 0.09 acre with a loss of 0.17 metric ton yield. Crop quantity and area were significantly correlated to each other. District Neelum shared 49% of the total crop loss, while 47% of the maize was raided at the altitudinal range of 2100-2500 m. crop raiding was highly significantly ( χ 2 = 1174.64 ; d f = 308 ;   p < 0.01 ) dependent upon distance to the forest. Linear regression revealed that maize quantity was determined by area, time and the total field area. Farmers faced 3.8 million PKRs loss due to crop damage by black bears. Despite the huge loss, the majority (23%) of the respondents did not respond to the query on mitigation measures indicating a poor adaptation of preventive measures. Preferred strategy to avoid crop damage was making noise (27.8%) when bears attacked their crops. A start of compensation scheme to the farmers is recommended that will have turned their negative attitude into a positive one toward the wildlife and black bear particularly. Study provides a new insight in human-bear conflict, particularly in spatial and temporal context of crop raiding in AJ&K.

Towards athermal organic-inorganic guided mode resonance filters.

We demonstrate guided-mode resonance filters featuring an amorphous TiO(2) layer fabricated by atomic layer deposition on a polymeric substrate. The thermal properties of such filters are studied in detail by taking into account both thermal expansion of the structure and thermo-optic coefficients of the materials. We show both theoretically and experimentally that these two effects partially compensate for each other, leading to nearly athermal devices. The wavelength shift of the resonance reflectance peak (< 1 nm) is a small fraction of the peak width (~11 nm) up to temperatures exceeding the room temperature by tens of degrees centigrade.

Food habits of indian crested porcupine (Hystrix indica) (Kerr 1792), in district Bagh, Azad Jammu and Kashmir.

The Indian Crested Porcupine (Hystrix indica) is classified as an agricultural pest species. It feeds on plants and crops; hence, it is responsible for massive financial losses worldwide. The current study was conducted to assess the diet composition of Indian Crested Porcupine in District Bagh, Azad Jammu and Kashmir (AJ&K). Thus, fecal samples were collected and examined from different sampling sites. Reference slides of the material collected from the study area were prepared for identification of dietary components in fecal pellets. A total of 80 fecal samples were collected and processed. Percent relative frequencies (P.R.F.) were calculated for each plant species recovered from pellets. Data revealed that Indian Crested Porcupine consumed 31 plant species in its diet, among them Zea mays (34.31±7.76) was the most frequently selected species followed by Rumex obtusifolius (15.32±2.57) and Melia azedarach (12.83±4.79). The study revealed that the greatest diversity of (n=20) plant species were consumed in summer season while minimum (n=13) species were used during winter. Among the parts of plants, stem was highly consumed in spring (57.2%) as compared to seed in fall (36.7%) while spikes and leaf were the least recovered parts from the fecal matter. The Berger-Parker diversity index showed highly diversified food (10.92) in the summer time of the year as compared to the autumn season (2.95). This study provides a baseline for the diet preference of this pest in the study area. Based on current findings, a detailed investigation on damage assessment, exploration, habitat use and management of Indian Crested Porcupine in AJ&K has been recommended.

Cannabidiol Ameliorates Cognitive Function via Regulation of IL-33 and TREM2 Upregulation in a Murine Model of Alzheimer's Disease.

There is a dire need for due innovative therapeutic modalities to improve outcomes of AD patients. In this study, we tested whether cannabidiol (CBD) improves outcomes in a translational model of familial AD and to investigate if CBD regulates interleukin (IL)-33 and triggering receptor expressed on myeloid cells 2 (TREM2), which are associated with improved cognitive function. CBD was administered to 5xFAD mice, which recapitulate early onset, familial AD. Behavioral tests and immunoassays were used to evaluate cognitive and motor outcomes. Our findings suggest that CBD treatment enhanced IL-33 and TREM2 expression, ameliorated the symptoms of AD, and retarded cognitive decline.

Food habits of indian crested porcupine (Hystrix indica) (Kerr 1792), in district Bagh, Azad Jammu and Kashmir / Hábitos alimentares do porco-espinho de crista indiano (Hystrix indica) (Kerr 1972) nos distritos de Bagh, Azad Jammu e Caxemira

The Indian Crested Porcupine (Hystrix indica) is classified as an agricultural pest species. It feeds on plants and crops; hence, it is responsible for massive financial losses worldwide. The current study was conducted to assess the diet composition of Indian Crested Porcupine in District Bagh, Azad Jammu and Kashmir (AJ&K). Thus, fecal samples were collected and examined from different sampling sites. Reference slides of the material collected from the study area were prepared for identification of dietary components in fecal pellets. A total of 80 fecal samples were collected and processed. Percent relative frequencies (P.R.F.) were calculated for each plant species recovered from pellets. Data revealed that Indian Crested Porcupine consumed 31 plant species in its diet, among them Zea mays (34.31±7.76) was the most frequently selected species followed by Rumex obtusifolius (15.32±2.57) and Melia azedarach (12.83±4.79). The study revealed that the greatest diversity of (n=20) plant species were consumed in summer season while minimum (n=13) species were used during winter. Among the parts of plants, stem was highly consumed in spring (57.2%) as compared to seed in fall (36.7%) while spikes and leaf were the least recovered parts from the fecal matter. The Berger-Parker diversity index showed highly diversified food (10.92) in the summer time of the year as compared to the autumn season (2.95). This study provides a baseline for the diet preference of this pest in the study area. Based on current findings, a detailed investigation on damage assessment, exploration, habitat use and management of Indian Crested Porcupine in AJ&K has been recommended.

O porco-espinho de crista indiano (Hystrix indica) é classificado como uma espécie de praga agrícola. Alimenta-se de plantas e colheitas; portanto, é responsável por enormes perdas financeiras em todo o mundo. O estudo atual foi realizado para avaliar a composição da dieta do porco-espinho de crista indiano nos distritos de Bagh, Azad Jammu e Caxemira (AJ&K). Assim, amostras fecais foram coletadas e examinadas em diferentes locais de amostragem. Lâminas de referência do material coletado na área de estudo foram preparadas para identificação dos componentes da dieta em pellets fecais. Um total de 80 amostras fecais foi coletado e processado. As frequências relativas percentuais (P.R.F.) foram calculadas para cada espécie de planta recuperada de pelotas. Os dados revelaram que o porco-espinho indiano consumiu 31 espécies de plantas em sua dieta, entre elas Zea mays (34,31 ± 7,76) foi a espécie mais selecionada, seguida por Rumex obtusifolius (15,32 ± 2,57) e Melia azedarach (12,83 ± 4,79). O estudo revelou que a maior diversidade de (n = 20) espécies de plantas foi consumida no verão, enquanto o mínimo (n = 13) espécies foi utilizado durante o inverno. Entre as partes das plantas, o caule foi muito consumido na primavera (57,2%) em relação à semente no outono (36,7%), enquanto a espiga e a folha foram as partes menos recuperadas da matéria fecal., O índice de diversidade de Berger-Parker mostrou alimentos altamente diversificados (10,92) no verão do ano em comparação com o outono (2,95). Este estudo fornece uma linha de base para a preferência alimentar dessa praga na área de estudo. Com base nas descobertas atuais, uma investigação detalhada sobre avaliação de danos, exploração, uso de habitat e gerenciamento de porco-espinho de crista indiano em AJ&K foi recomendada.

Food habits of indian crested porcupine (Hystrix indica) (Kerr 1792), in district Bagh, Azad Jammu and Kashmir

Abstract The Indian Crested Porcupine (Hystrix indica) is classified as an agricultural pest species. It feeds on plants and crops; hence, it is responsible for massive financial losses worldwide. The current study was conducted to assess the diet composition of Indian Crested Porcupine in District Bagh, Azad Jammu and Kashmir (AJ&K). Thus, fecal samples were collected and examined from different sampling sites. Reference slides of the material collected from the study area were prepared for identification of dietary components in fecal pellets. A total of 80 fecal samples were collected and processed. Percent relative frequencies (P.R.F.) were calculated for each plant species recovered from pellets. Data revealed that Indian Crested Porcupine consumed 31 plant species in its diet, among them Zea mays (34.31±7.76) was the most frequently selected species followed by Rumex obtusifolius (15.32±2.57) and Melia azedarach (12.83±4.79). The study revealed that the greatest diversity of (n=20) plant species were consumed in summer season while minimum (n=13) species were used during winter. Among the parts of plants, stem was highly consumed in spring (57.2%) as compared to seed in fall (36.7%) while spikes and leaf were the least recovered parts from the fecal matter. The Berger-Parker diversity index showed highly diversified food (10.92) in the summer time of the year as compared to the autumn season (2.95). This study provides a baseline for the diet preference of this pest in the study area. Based on current findings, a detailed investigation on damage assessment, exploration, habitat use and management of Indian Crested Porcupine in AJ&K has been recommended.

Resumo O porco-espinho de crista indiano (Hystrix indica) é classificado como uma espécie de praga agrícola. Alimenta-se de plantas e colheitas; portanto, é responsável por enormes perdas financeiras em todo o mundo. O estudo atual foi realizado para avaliar a composição da dieta do porco-espinho de crista indiano nos distritos de Bagh, Azad Jammu e Caxemira (AJ&K). Assim, amostras fecais foram coletadas e examinadas em diferentes locais de amostragem. Lâminas de referência do material coletado na área de estudo foram preparadas para identificação dos componentes da dieta em pellets fecais. Um total de 80 amostras fecais foi coletado e processado. As frequências relativas percentuais (P.R.F.) foram calculadas para cada espécie de planta recuperada de pelotas. Os dados revelaram que o porco-espinho indiano consumiu 31 espécies de plantas em sua dieta, entre elas Zea mays (34,31 ± 7,76) foi a espécie mais selecionada, seguida por Rumex obtusifolius (15,32 ± 2,57) e Melia azedarach (12,83 ± 4,79). O estudo revelou que a maior diversidade de (n = 20) espécies de plantas foi consumida no verão, enquanto o mínimo (n = 13) espécies foi utilizado durante o inverno. Entre as partes das plantas, o caule foi muito consumido na primavera (57,2%) em relação à semente no outono (36,7%), enquanto a espiga e a folha foram as partes menos recuperadas da matéria fecal., O índice de diversidade de Berger-Parker mostrou alimentos altamente diversificados (10,92) no verão do ano em comparação com o outono (2,95). Este estudo fornece uma linha de base para a preferência alimentar dessa praga na área de estudo. Com base nas descobertas atuais, uma investigação detalhada sobre avaliação de danos, exploração, uso de habitat e gerenciamento de porco-espinho de crista indiano em AJ&K foi recomendada.

Food habits of indian crested porcupine (Hystrix indica) (Kerr 1792), in district Bagh, Azad Jammu and Kashmir / Hábitos alimentares do porco-espinho de crista indiano (Hystrix indica) (Kerr 1972) nos distritos de Bagh, Azad Jammu e Caxemira

The Indian Crested Porcupine (Hystrix indica) is classified as an agricultural pest species. It feeds on plants and crops; hence, it is responsible for massive financial losses worldwide. The current study was conducted to assess the diet composition of Indian Crested Porcupine in District Bagh, Azad Jammu and Kashmir (AJ&K). Thus, fecal samples were collected and examined from different sampling sites. Reference slides of the material collected from the study area were prepared for identification of dietary components in fecal pellets. A total of 80 fecal samples were collected and processed. Percent relative frequencies (P.R.F.) were calculated for each plant species recovered from pellets. Data revealed that Indian Crested Porcupine consumed 31 plant species in its diet, among them Zea mays (34.31±7.76) was the most frequently selected species followed by Rumex obtusifolius (15.32±2.57) and Melia azedarach (12.83±4.79). The study revealed that the greatest diversity of (n=20) plant species were consumed in summer season while minimum (n=13) species were used during winter. Among the parts of plants, stem was highly consumed in spring (57.2%) as compared to seed in fall (36.7%) while spikes and leaf were the least recovered parts from the fecal matter. The Berger-Parker diversity index showed highly diversified food (10.92) in the summer time of the year as compared to the autumn season (2.95). This study provides a baseline for the diet preference of this pest in the study area. Based on current findings, a detailed investigation on damage assessment, exploration, habitat use and management of Indian Crested Porcupine in AJ&K has been recommended.

O porco-espinho de crista indiano (Hystrix indica) é classificado como uma espécie de praga agrícola. Alimenta-se de plantas e colheitas; portanto, é responsável por enormes perdas financeiras em todo o mundo. O estudo atual foi realizado para avaliar a composição da dieta do porco-espinho de crista indiano nos distritos de Bagh, Azad Jammu e Caxemira (AJ&K). Assim, amostras fecais foram coletadas e examinadas em diferentes locais de amostragem. Lâminas de referência do material coletado na área de estudo foram preparadas para identificação dos componentes da dieta em pellets fecais. Um total de 80 amostras fecais foi coletado e processado. As frequências relativas percentuais (P.R.F.) foram calculadas para cada espécie de planta recuperada de pelotas. Os dados revelaram que o porco-espinho indiano consumiu 31 espécies de plantas em sua dieta, entre elas Zea mays (34,31 ± 7,76) foi a espécie mais selecionada, seguida por Rumex obtusifolius (15,32 ± 2,57) e Melia azedarach (12,83 ± 4,79). O estudo revelou que a maior diversidade de (n = 20) espécies de plantas foi consumida no verão, enquanto o mínimo (n = 13) espécies foi utilizado durante o inverno. Entre as partes das plantas, o caule foi muito consumido na primavera (57,2%) em relação à semente no outono (36,7%), enquanto a espiga e a folha foram as partes menos recuperadas da matéria fecal., O índice de diversidade de Berger-Parker mostrou alimentos altamente diversificados (10,92) no verão do ano em comparação com o outono (2,95). Este estudo fornece uma linha de base para a preferência alimentar dessa praga na área de estudo. Com base nas descobertas atuais, uma investigação detalhada sobre avaliação de danos, exploração, uso de habitat e gerenciamento de porco-espinho de crista indiano em AJ&K foi recomendada.

An improved method for evaluation of blood coagulation in heparinized blood.

A simple method designed to allow coagulation studies in heparinized blood was evaluated. Citrated plasma containing heparin was incubated with an anion-exchange resin commercially available in tablet form (Heparsorb). After centrifugation of the mixture, coagulation studies were carried out on the supernatant plasma. One heparin neutralizer (HN) tablet was capable of removing as much as 43 units of heparin from 1 ml plasma within 10 min of incubation. When normal plasma was exposed to the heparin neutralizer, little or no change in the activities of clotting factors XII, XI, VIII, VII, X, V, and II was observed. However, the resin caused a substantial (44%) loss of factor IX-activity from normal plasma. This loss of factor IX-activity was not observed with plasma from patients receiving coumadin therapy. The results of coagulation tests (aPTT, PT, TT) and determination of fibrinogen/fibrin degradation products performed with HN-treated plasmas from nine patients receiving heparin therapy for thromboembolic disease and seven patients undergoing cardiopulmonary bypass operations were virtually identical to those obtained for the same persons before heparin was administered. The method, which is suitable for the routine clinical laboratory, may be useful in the hemostatic evaluation of critically ill patients who experience bleeding complications while receiving heparin therapy, in the laboratory control of coumadin therapy during the heparin-coumadin overlap period, and in the rapid identification of heparin contamination of blood specimens as a cause of an unexplained aPTT prolongation.

Purification and characterization of hemolysin from Porphyromonas gingivalis A7436.

Porphyromonas gingivalis, a periodontal pathogen, has the ability to lyse erythrocytes. The hemolytic activity of P. gingivalis A7436 was purified as a 45-kDa protein from the culture supernatant of a 3-days old culture using nickel-nitrilotriacetic acid chromatography. Erythrocytes treated with purified P. gingivalis hemolysin showed the presence of pores and extracellular debris by scanning electron microscopy. Active immunization of mice with 15 micrograms hemolysin induced neutralizing antibodies to hemolysin. Heating at 60 degrees C and treatment with trypsin and dithiothreitol abolished hemolytic activity, while incubation with the protease inhibitor Na-p-tosyl-L-lysine chloromethyl ketone caused no effect. We report here for the first time purification of a hemolysin from P. gingivalis A7436. The amino acid sequence of an internal peptide of hemolysin showed sequence similarity with fimbrillin from P. gingivalis HG564. However, the amino acid composition of purified hemolysin was different from that of P. gingivalis fimbrillin. Also, the ability to lyse but not agglutinate erythrocytes and to bind to nickel-nitrilotriacetic acid differentiates P. gingivalis hemolysin from fimbrillin.

Purification and partial characterisation of a novel 66-kDa seroreactive protein of Mycobacterium tuberculosis H37Rv.

A seroreactive protein (TB66) was purified from culture filtrate (CF) and cell sonicate (CS) of Mycobacterium tuberculosis H37Rv by immobilised metal affinity chromatography (IMAC) on a Ni-nitrilotriacetic acid (NTA) column. The TB66 preparations obtained by IMAC contained predominantly a 66-kDa protein with a pI of c. 5.5 as determined by two-dimensional electrophoresis. TB66 was detected in the CF as early as 1 week of growth of H37Rv. The NH2-terminal amino-acid sequence showed 85% homology with the N-terminal sequence of bovine serum albumin (BSA) and 80% homology with human serum albumin. Amino-acid analysis indicated a difference in the amino-acid content of TB66 when compared to BSA, with an abundance of acidic amino acids. A monoclonal antibody (MAb) OD4AG3, raised in this laboratory against an M. avium complex (MAC 101) sonicate cross-reacting with H37Rv sonicate, recognised a heat-stable and trypsin-sensitive epitope of this protein. TB66 was also recognised by MAbs IT1 and IT20 which also react with the 14-kDa antigen of the M. tuberculosis complex. Antibodies against TB66 were present in the sera of 62 of 64 patients with tuberculosis; sera from normal healthy individuals showed no significant reactivity. TB66 appears to be a predominant secretory protein of M. tuberculosis and could play an important role in the pathogenesis of this organism.

Skin reactivity and fibronectin-binding property of TB66 (66-kDa protein of Mycobacterium tuberculosis).

A 66-kDa protein (TB66) was purified from culture filtrate (CF) and cell sonicate (CS) of Mycobacterium tuberculosis H37Rv by immobilised metal affinity chromatography (IMAC) on a Ni-nitrilotriacetic acid (NTA) column. TB66 was found to be a fibronectin-binding protein as determined by ELISA and could be purified by affinity chromatography with fibronectin-Sepharose. A similar 66-kDa protein could be isolated also from M. bovis, M. bovis BCG, M. africanum and M. tuberculosis H37Ra by IMAC, but not from any other mycobacteria. The NH2-terminal amino-acid sequence of TB66 from H37Rv and M. bovis was identical and showed 85% homology with the N-terminal sequence of bovine serum albumin (BSA). A monoclonal antibody (MAb) OD4AG3 recognised a heat-stable and trypsin-sensitive epitope near the C-terminal end of TB66. This MAb also recognised the 66-kDa protein isolated from the other members of the M. tuberculosis complex. In tests of immunogenicity, TB66 elicited a delayed type hypersensitivity reaction in guinea-pigs immunised with either TB66 or with M. tuberculosis H37Rv. TB66 also elicited an antibody response in immunised guinea-pigs and stimulated murine macrophages to produce tumour necrosis factor.

Isolation of a contact-dependent haemolysin from Mycobacterium tuberculosis.

Contact-dependent haemolytic activity was observed with cells of Mycobacterium tuberculosis H(37)Rv and M. tuberculosis H(37)Ra, but not with those of M. bovis, M. bovis BCG and M. africanum. Culture filtrates of all these strains did not exhibit any haemolytic activity. M. tuberculosis H(37)Rv was subsequently used for the isolation of haemolysin. Haemolytic activity was retained in the cell debris even after sonication of the cells and treatment with Tween 80 and lysozyme. Solubilisation of haemolysin was possible only after the cell debris was washed with ethanol 70% and then treated with Tween 80 0.1%. The haemolysin thus obtained showed a micellar M(r) of >200000 by gel-filtration on Sephadex G-200 and a subunit M(r) of 66000 by SDS-PAGE. It was sensitive to trypsin but stable when heated at 60 degrees C for 10 min. Polyclonal serum raised in rabbits against the haemolysin neutralised the haemolytic activity. The N-terminal amino-acid sequence of the 66-kDa subunit of haemolysin showed identity with TB66, the 66-kDa secretory protein of M. tuberculosis, and 30% homology with the haemolysin A precursor of Vibrio cholerae. Phosphatidylglycerol inhibited lysis of sheep erythrocytes by the haemolysin and is probably the receptor for the haemolysin. Haemolysin not only lysed erythrocytes, but was also cytotoxic to human lung cells. It appears that, among the members of the M. tuberculosis complex, the cell-bound contact-dependent haemolysin/cytolysin is restricted to M. tuberculosis and it may be associated with the pathogenesis of M. tuberculosis.

Haemolysin from Mycobacterium avium complex isolates from AIDS patients.

Cell-bound haemolytic activity was observed in isolates of Mycobacterium avium complex (MAC) from AIDS patients. M. avium type strains showed negligible activity. None of the culture supernates exhibited any haemolytic activity. Zwitterionic detergent 3-[(3-cholamidopropyl)-dimethyl-ammonio]-1-propanesulphonate (CHAPS) was used to extract haemolysin from ethanol-treated M. avium complex strain 101 (MAC101) cells. Haemolysin was isolated from CHAPS extract (CE) by metal affinity chromatography and identified as a 32-kDa protein by polyclonal antibodies raised against M. tuberculosis haemolysin. Treatment of CE with trypsin resulted in reduction of haemolytic activity, whereas heating at 100 degrees C for 10 min did not affect its activity. A similar 32-kDa haemolysin was extracted from cells of M. avium K128 which was isolated from a monkey infected with simian immunodeficiency virus (SIV). The haemolysin produced by M. avium strains isolated from AIDS patients may be associated with the pathogenesis of M. avium infection.

Immunoaffinity chromatographic isolation of a high molecular weight seroreactive protein from Mycobacterium leprae cell sonicate.

The purpose of this study was to isolate Mycobacterium leprae antigen(s) by immunoaffinity chromatography using immunoglobulins from leprosy patients and from rabbit anti-M. leprae hyperimmune serum coupled to CNBr-Sepharose 4B. A high molecular weight (M(r)) M. leprae protein (MLP) with a subunit M(r) of 22,000 was isolated. MLP was recognized by monoclonal antibody MMPII1G4 which is known to react with MMPII, a 22 kDa protein of M. leprae. The N-terminal sequence of the 22 kDa subunit (Met-gln-gly-asp-pro-asp-val-leu-arg-leu-leu-asn-glu-gln-leu-thr) was identical to MMPII and to antigen D (bacterioferritin) of M. paratuberculosis. It showed 44% homology with N-terminal end of E. coli bacterioferritin. In ELISA, MLP showed 100% and 60% positivity with leprosy and TB sera respectively as compared to normal healthy sera. The role of bacterioferritin in M. leprae and the importance of MLP as an immunogen has been discussed.

The foster care system in India.

The author has undertaken the difficult task of giving us an overall view of the foster care picture of all India, the first such panorama we have received.

Comparative antigenic analysis of Mycobacterium avium complex (MAC) isolates from AIDS patients.

Sonicates of several Mycobacterium avium complex (MAC) strains isolated from acquired immunodeficiency syndrome (AIDS) patients were characterized in order to study the prominent antigens of these strains. Sonicates of 6-week-old cultures were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting. A major 12 kDa glycoprotein antigen was observed in all the sonicates along with other proteins ranging up to 100 kDa. Western blotting, using the 12 kDa M. leprae 'specific' murine monoclonal antibody (MAb) MLO6, indicated the presence of a determinant in the 12 kDa antigen (in all the MAC isolates studied) which was immunologically cross-reactive with the 12 kDa antigen of M. leprae. The transparent variant of MAC 101 also demonstrated MLO6 reactivity while the opaque variant did not. Polyclonal antiserum raised against MAC 101 sonicate reacted with all the MAC isolates in immunodiffusion. These observations point to the cross-reactivity between these strains and suggest that they possess a M. leprae 'specific' determinant on a cross-reacting component which could be involved in virulence.

Immunological evaluation of a 12-kilodalton protein of Mycobacterium tuberculosis by enzyme-linked immunosorbent assay.

OBJECTIVE: To purify and study the seroreactivity of native and recombinant 12-kilodalton protein of Mycobacterium tuberculosis H37Rv. DESIGN: M. tuberculosis H37Rv cells and Escherichia coli XL-1 containing the plasmid PRL4 encoding the M. tuberculosis heat shock protein GroES homolog were used as sources for the purification of native and recombinant 12 kD of M. tuberculosis respectively. The seroreactivity of the 12 kDs was studied by ELISA using sera from 35 leprosy and 25 active pulmonary tuberculosis (TB) patients, and from 10 normal healthy controls. RESULTS: The 12 kD protein was purified from H37Rv extract (s12 kD) and from recombinant E. coli (r12 kD) by ultrafiltration and MonoQ fast pressure liquid chromatography (FPLC). Analysis of s12 kD and r12 kD by SDS-PAGE revealed a single protein band in both cases with an approximate molecular weight of 12,000 which was recognized by monoclonal antibody SA-12 in immunoblotting. Both the proteins exhibited a pI of approximately 4.6 by isoelectric focusing. Both the 12 kD proteins exhibited 96% positivity with TB sera as compared to normal control sera (P < 0.01). Only one serum sample from the 35 leprosy sera tested exhibited binding to both the s12 kD and r12 kD proteins. Delayed type hypersensitivity reaction to the 12 kD proteins was elicited in guinea pigs that had been immunized with H37Rv sonicate. CONCLUSION: The 12 kD protein could be easily purified and could serve as a valuable serodiagnostic tool in the screening of TB cases from a large population in an endemic area.