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In this paper, a new method for pesticide detection based on porous silicon microcavities was proposed. First, the aptamer of acetamiprid was fixed in the functionalized porous silicon microcavity. Then, the complementary strand of the aptamer modified with graphene quantum dots was hybridized with the aptamer, and the effective refractive index of the porous silicon microcavity increased. When acetamiprid was added, it bound to the aptamer due to its stronger specificity. The aptamer separated from the complementary strand of the coupled graphene quantum dots and bound to acetamiprid, resulting in a decrease in the effective refractive index, and its central wavelength of reflection spectrum was blue shifted, and the blueshift increased with the increase of acetamiprid concentration. The relationship between the change in central wavelength and the concentration of acetamiprid was analysed, and the detection limit of acetamiprid was 151 nm.

In this paper, a grayscale variation of fluorescence Image based on porous silicon (PSi) Bragg mirrors (Bm) was proposed to detect pesticides quickly and easily. In this way, CdSe/ZnS quantum dots (QDs) were used as fluorescence labelling to weaken the fluorescence intensity, and a digital imaging method was used for detection. The QDs were coupled with complementary strand aptamers and interacted with aptamers immobilized on the PSi‘inner wall; thus, the fluorescence of the QDs produced by the PSi Bm was further enhanced. Acetamiprid would bind specifically to its corresponding aptamer, the complementary strand aptamers of the coupled QD would be replaced, and the fluorescence intensity would weaken. The fluorescence image of the PSi Bm surface was acquired by imaging device, and the acetamiprid pesticide was detected by computing the average grey value variation of the image. Experimental results presented that the variation in the average grey value increased with increasing pesticide concentration and presented a linear relationship within a certain range. The detection limit of acetamiprid pesticide with this type of way was 2.8 nM, indicating that this method achieved a low-cost, rapid and simple detection of pesticide.

Bovine leukemia virus (BLV) is an enveloped virus, found worldwide that can infect cattle and induce many subclinical symptoms and malignant tumors. BLV infection causes severe economic losses in the cattle industry. The identification of BLV-infected cattle for segregation or elimination would be the most effective way to halt the spread of BLV infection on farms, owing to the lack of effective treatments and vaccines. Therefore, antibody detection against the viral glycoprotein gp51 is an effective method for diagnosing BLV-infected animals. In this study, ten different subregions of gp51 containing a common B cell epitope are vital for developing antigens as epitope-driven vaccine design and immunological assays. Such antigens were produced in Escherichia coli expression system to react with antibodies in the serum from BLV-infected cattle and compete for antigenicity. Recombinant His-gp51

In road construction, a large number of excavated soils need to be treated with stabilizers. The addition of superabsorbent polymer (SAP) can improve the road performance of these stabilized soils. In order to predict roadbed deformation, dynamic triaxial tests were carried out on cemented soil containing SAP to investigate its resilient and plastic strain behavior. The effects of SAP content, cyclic stress ratio, and loading frequency on cement-stabilized soils with SAP were analyzed combined with the number of cycles. This study demonstrates how these influencing factors effect the resilient strain, dynamic elastic modulus, and accumulated plastic strain, which are crucial to better understanding the strain behavior of cement-stabilized soil with SAP. The results show that SAP can significantly improve the brittle failure characteristics and dynamic strength of cement-stabilized soil. Soil with higher SAP content possesses smaller accumulated plastic strain; with the increase in the cyclic stress ratio, the dynamic elastic modulus decreases significantly, whereas the accumulated plastic strain has the opposite trend. In addition, the lower frequency produces larger cumulative axial strain.

In this work, a new dual signal light detection method based on porous silicon Bragg mirror (PSBM) and biological labelling with quantum dots (QDs) is proposed for the detection of beta-lactoglobulin (β-lg). The first signal light is a probe light emitted by a laser with wavelength of 633 nm, which enters the PSBM and is reflected from the surface. The wavelength of the probe light is located at the edge of the PSBM band gap, where it has the lowest reflectivity. β-lg antibodies is labelled with CdSe/ZnS QDs and reacts with β-lg molecules have been fixed to the inner wall of the porous silicon pores. Due to the specific binding of biomolecules in PSBM, the refractive index of the device increases, resulting in the enhancement of detection reflected light. The QDs play the role of refractive index amplification. The second signal light is the fluorescence of QDs in immune reactants. QDs produce fluorescence at 630 nm when excited by a short-wavelength laser. The fluorescence signal is further enhanced by PSBM. The superimposed images of two kinds of light on the surface of PSBM are obtained by digital microscope at the same time. By calculating the average grey value change of the image before and after biological reaction, β-lg can be detected with high sensitivity. The detection limit of β-lg was 0.12 ng/mL. The experimental results showed that the PSBM-based dual signal light method could be used to detect the content of cow milk adulterated in β-lg free camel milk.

Background Bovine leukemia virus (BLV) causes enzootic bovine leukosis, the most common neoplastic disease of cattle. Previously, we reported the luminescence syncytium induction assay (LuSIA), an assay for BLV infectivity based on CC81-BLU3G cells, which form syncytia expressing enhanced green fluorescent protein (EGFP) when co-cultured with BLV-infected cells. To develop a more sensitive LuSIA, we here focused on the glucocorticoid response element (GRE) within the U3 region of the BLV long terminal repeat (LTR). Methods We changed five nucleotide sites of the GRE in a pBLU3-EGFP reporter plasmid containing the BLV-LTR U3 region promoter by site-directed mutagenesis and we then constructed a new reporter plasmid (pBLU3GREM-EGFP) in which the EGFP reporter gene was expressed under control of the GRE-mutated LTR-U3 promoter. We also established a new CC81-derived reporter cell line harboring the GRE-mutated LTR-U3 promoter (CC81-GREMG). To evaluate the sensibility, the utility and the specificity of the LuSIA using CC81-GREMG, we co-cultured CC81-GREMG cells with BLV-persistently infected cells, free-viruses, white blood cells (WBCs) from BLV-infected cows, and bovine immunodeficiency-like virus (BIV)- and bovine foamy virus (BFV)-infected cells. Results We successfully constructed a new reporter plasmid harboring a mutation in the GRE and established a new reporter cell line, CC81-GREMG; this line was stably transfected with pBLU3GREM-EGFP in which the EGFP gene is expressed under control of the GRE-mutated LTR-U3 promoter and enabled direct visualization of BLV infectivity. The new LuSIA protocol using CC81-GREMG cells measures cell-to-cell infectivity and cell-free infectivity of BLV more sensitively than previous protocol using CC81-BLU3G. Furthermore, it did not respond to BIV and BFV infections, indicating that the LuSIA based on CC81-GREMG is specific for BLV infectivity. Moreover, we confirmed the utility of a new LuSIA based on CC81-GREMG cells using white blood cells (WBCs) from BLV-infected cows. Finally, the assay was useful for assessing the activity of neutralizing antibodies in plasma collected from BLV-infected cows. Conclusion The new LuSIA protocol is quantitative and more sensitive than the previous assay based on CC81-BLU3G cells and should facilitate development of several new BLV assays.

Perinatal transmission plays a critical role in the spread of bovine leukemia virus (BLV) infection in cattle herds. In the Holstein breed, we previously identified BLV resistant and susceptible bovine leukocyte antigen (BoLA)-DRB3 alleles, including BoLA-DRB3*009:02 and *014:01:01 with a low BLV proviral load (PVL), and *015:01 and *012:01 with a high PVL. Here, we evaluated the perinatal BLV transmission risk in dams with different BoLA-DRB3 alleles. BoLA-DRB3 alleles of 120 dam-calf pairs from five dairy farms in Japan were identified, their PVL was quantified using the BLV-Coordination of Common Motifs (CoCoMo)-qPCR-2 assay. Ninety-six dams were BLV-positive, and 29 gave birth to BLV-infected calves. Perinatal transmission frequency was 19% in dams with resistant alleles suppressed to a low PVL level, and 38% and 25% in dams with susceptible and neutral alleles that maintained high PVL levels, respectively. Notably, all calves with resistant alleles were BLV free, whereas 30% of calves with susceptible genes were infected. Thus, vertical transmission risk was extremely lower for dams and calves with resistant alleles compared to those with susceptible alleles. Our results can inform the development of effective BLV eradication programs under field conditions by providing necessary data to allow for optimal selection of dams for breeding.

Bovine leukemia virus (BLV) causes enzootic bovine leukosis, a malignant form of B-cell lymphoma, and is closely related to human T-cell leukemia viruses. We investigated whether BLV infection affects host genes associated with DNA mismatch repair (MMR). Next-generation sequencing of blood samples from five calves experimentally infected with BLV revealed the highest expression levels of seven MMR genes (EXO1, UNG, PCNA, MSH2, MSH3, MSH6, and PMS2) at the point of peak proviral loads (PVLs). Furthermore, MMR gene expression was only upregulated in cattle with higher PVLs. In particular, the expression levels of MSH2, MSH3, and UNG positively correlated with PVL in vivo. The expression levels of all seven MMR genes in pig kidney-15 cells and the levels of PMS2 and EXO1 in HeLa cells also increased tendencies after transient transfection with a BLV infectious clone. Moreover, MMR gene expression levels were significantly higher in BLV-expressing cell lines compared with those in the respective parental cell lines. Expression levels of MSH2 and EXO1 in BLV-infected cattle with lymphoma were significantly lower and higher, respectively, compared with those in infected cattle in vivo. These results reveal that BLV infection affects MMR gene expression, offering new candidate markers for lymphoma diagnosis.

Bovine leukemia virus (BLV) causes enzootic bovine leukosis, the most common neoplastic disease in cattle. We previously reported the development and protocol of the luminescence syncytium induction assay (LuSIA), a method for evaluating BLV infectivity based on CC81-GREMG cells. These cells form syncytia expressing enhanced green fluorescent protein when co-cultured with BLV-infected cells. Recently, we confirmed CAT1/SLC7A1 functions as a receptor of BLV. Here, we focused on CAT1/SLC7A1 to increase the sensitivity of LuSIA. We constructed a bovine CAT1-expressing plasmid and established a new CC81-GREMG-derived reporter cell line highly expressing bovine CAT1 (CC81-GREMG-CAT1). The new LuSIA protocol using CC81-GREMG-CAT1 cells measures cell-to-cell infectivity and cell-free infectivity of BLV faster and with greater sensitivity than the previous protocol using CC81-GREMG. The new LuSIA protocol is quantitative and more sensitive than the previous assay based on CC81-GREMG cells and will facilitate the development of several new BLV assays.

The cutter head parts of the tunnel boring machine (TBM) are of different sizes, and the cutting platform is of large size. The panoramic view of the cutting platform that meets the accuracy requirements through image stitching is a prerequisite for realizing the visual guidance of autonomous cutting. An image stitching algorithm is studied to reduce the complexity of the algorithm on the basis of ensuring the stitching precision. The ORB algorithm is selected to implement feature detection, and a series of strategies are adopted on this basis. The ROI region is extracted, and only the images in the region of interest are matched. Then, the feature detection, description and matching are performed by the ORB feature detection algorithm and Hamming distance. The feature points are purified by the RANSAC algorithm, and the transformation matrix H is calculated. Finally, the image fusion is carried out. The experimental results show that the ROI-based ORB image stitching algorithm satisfies the accuracy requirements of subsequent visual guidance.

Background Bovine leukemia virus (BLV), which is closely related to human T-cell leukemia virus, is the etiological agent of enzootic bovine leukosis, a disease characterized by a highly prolonged course involving persistent lymphocytosis and B-cell lymphoma. The bovine major histocompatibility complex class II region plays a key role in the subclinical progression of BLV infection. In this study, we aimed to evaluate the roles of CD4+ T-cell epitopes in disease progression in cattle. Methods We examined five Japanese Black cattle, including three disease-susceptible animals, one disease-resistant animal, and one normal animal, classified according to genotyping of bovine leukocyte antigen (BoLA)-DRB3 and BoLA-DQA1 alleles using polymerase chain reaction sequence-based typing methods. All cattle were inoculated with BLV-infected blood collected from BLV experimentally infected cattle and then subjected to CD4+ T-cell epitope mapping by cell proliferation assays. Results Five Japanese Black cattle were successfully infected with BLV, and CD4+ T-cell epitope mapping was then conducted. Disease-resistant and normal cattle showed low and moderate proviral loads and harbored six or five types of CD4+ T-cell epitopes, respectively. In contrast, the one of three disease-susceptible cattle with the highest proviral load did not harbor CD4+ T-cell epitopes, and two of three other cattle with high proviral loads each had only one epitope. Thus, the CD4+ T-cell epitope repertoire was less frequent in disease-susceptible cattle than in other cattle. Conclusion Although only a few cattle were included in this study, our results showed that CD4+ T-cell epitopes may be associated with BoLA-DRB3-DQA1 haplotypes, which conferred differential susceptibilities to BLV proviral loads. These CD4+ T-cell epitopes could be useful for the design of anti-BLV vaccines targeting disease-susceptible Japanese Black cattle. Further studies of CD4+ T-cell epitopes in other breeds and using larger numbers of cattle with differential susceptibilities are required to confirm these findings.

Bovine leukemia virus (BLV) infects cattle and causes serious problems for the cattle industry, worldwide. Vertical transmission of BLV occurs via in utero infection and ingestion of infected milk and colostrum. The aim of this study was to clarify whether milk is a risk factor in BLV transmission by quantifying proviral loads in milk and visualizing the infectivity of milk. We collected blood and milk from 48 dams (46 BLV seropositive dams and 2 seronegative dams) from seven farms in Japan and detected the BLV provirus in 43 blood samples (89.6%) but only 22 milk samples (45.8%) using BLV-CoCoMo-qPCR-2. Although the proviral loads in the milk tended to be lower, a positive correlation was firstly found between the proviral loads with blood and milk. Furthermore, the infectivity of milk cells with BLV was visualized ex vivo using a luminescence syncytium induction assay (LuSIA) based on CC81-GREMG cells, which form syncytia expressing enhanced green fluorescent protein (EGFP) in response to BLV Tax and Env expressions when co-cultured with BLV-infected cells. Interestingly, in addition to one BLV-infected dam with lymphoma, syncytia with EGFP fluorescence were observed in milk cells from six BLV-infected, but healthy, dams by an improved LuSIA, which was optimized for milk cells. This is the first report demonstrating the infectious capacity of cells in milk from BLV-infected dams by visualization of BLV infection ex vivo. Thus, our results suggest that milk is a potential risk factor for BLV vertical spread through cell to cell transmission.

Bovine leukemia virus (BLV) is the causative agent of enzootic bovine leukosis. Polymorphism in bovine lymphocyte antigen (BoLA)-DRB3 alleles is related to susceptibility to BLV proviral load (PVL), which is a useful index for estimating disease progression and transmission risk. However, whether differential BoLA-DRB3 affects BLV infectivity remains unknown. In a three-year follow-up investigation using a luminescence syncytium induction assay for evaluating BLV infectivity, we visualized and evaluated the kinetics of BLV infectivity in cattle with susceptible, resistant and neutral BoLA-DRB3 alleles which were selected from 179 cattle. Susceptible cattle showed stronger BLV infectivity than both resistant and neutral cattle. The order of intensity of BLV infectivity was as follows: susceptible cattle &gt, neutral cattle &gt, resistant cattle. BLV infectivity showed strong positive correlation with PVL at each testing point. BLV-infected susceptible cattle were found to be at higher risk of horizontal transmission, as they had strong infectivity and high PVL, whereas BLV-infected resistant cattle were low risk of BLV transmission owing to weak BLV infection and low PVL. Thus, this is the first study to demonstrate that the BoLA-DRB3 polymorphism is associated with BLV infection.

Bovine leukemia virus (BLV) is the causative agent of enzootic bovine leukosis, the most common neoplastic disease of cattle, which is closely related to human T-cell leukemia viruses. BLV has spread worldwide and causes a serious problem for the cattle industry. The cellular receptor specifically binds with viral envelope glycoprotein (Env) and this attachment mediates cell fusion to lead virus entry. BLV Env reportedly binds to cationic amino acid transporter 1 (CAT1)/SLC7A1, but whether the CAT1/SLC7A1 is an actual receptor for BLV remains unknown. Here, we showed that CAT1 functioned as an infection receptor, interacting with BLV particles. Cells expressing undetectable CAT1 levels were resistant to BLV infection but became highly susceptible upon CAT1 overexpression. CAT1 exhibited specific binding to BLV particles on the cell surface and co-localized with the Env in endomembrane compartments and membrane. Knockdown of CAT1 in permissive cells significantly reduced binding to BLV particles and BLV infection. In addition, bovine serum with neutralizing activity from a BLV-infected cattle inhibited BLV particles. Expression of CAT1 from various species demonstrated no species-specificity for BLV infection, implicating CAT1 as a functional BLV receptor responsible for its broad host range. These findings provide insights for BLV infection and for developing new strategies for treating BLV and preventing its spread.Author SummaryBovine leukemia virus (BLV), which can infect a variety of animal species and induce lymphoma in cattle, is a member of the familyRetroviridae. BLV induces huge economic losses by not only lymphoma but also subclinical forms of the disease. In addition, BLV is frequently used as an animal model of human T-cell leukemia virus (HTLV), as BLV has many similar characteristics to HTLV. Thus, understanding BLV pathogenesis contribute to resolve not only BLV-but also HTLV-induced problems. Retroviral envelope glycoprotein (Env) is specifically recognized by the cellular receptor at cell surface, which induces a conformational changes between viral and cell membrane to entry. Thus, the elucidation of cellular receptor for BLV infection is very important for virus entry. However, the BLV receptor has not been identified yet. In the current study, we found that BLV Env protein binds to cationic amino acid transporter 1 (CAT1)/SLC7A1 at cell surface, artificial expression of CAT1 in CAT1-negative cells confers the cells susceptible to BLV infection, and CAT1-silencing significantly reduces BLV infection, concluding that CAT1 is the BLV receptor. These findings will have far reaching great advantages of insights in the retrovirus study.

Epigallocatechin gallate (EGCG) is the major polyphenolic compound of green tea. Polyphenolic compounds were extracted from the leaf of Camellia sinensis (Japanese green tea), and the minimum inhibitory concentration against canine oral bacteria was measured. Subsequently, we investigated the inhibitory effects of polyphenolic compounds and EGCG on the growth of canine oral bacteria. EGCG showed antimicrobial activity against a model bacterium, Streptococcus mutans. Our results indicate that EGCG can inhibit the growth and biofilm formation of S. mutans and that EGCG does not interact with streptococcal lipoteichoic acid (LTA). Furthermore, our findings suggest that EGCG interacts with other component(s) of the bacterial membrane aside from streptococcal LTA to inhibit biofilm formation and damage biofilms.

Bovine leukemia virus (BLV) is associated with enzootic bovine leukosis and is closely related to human T cell leukemia virus (HTLV). The cytotoxic T lymphocyte (CTL) plays a key role in suppressing the progression of disease caused by BLV. T and B cell epitopes in BLV have been studied, but CD8(+) CTL epitopes remain poorly understood. We used a library of 115 synthetic peptides covering the entirety of the Env proteins (gp51 and gp30), the Gag proteins (p15, p24, and p12), and the Tax protein of BLV to identify 11 novel CD8(+) T cell epitopes (gp51N5, gp51N11, gp51N12, gp30N5, gp30N6, gp30N8, gp30N16, tax16, tax18, tax19, and tax20) in four calves experimentally infected with BLV. The number of CD8(+) T cell epitopes that could be identified in each calf correlated with the BLV proviral load. Interestingly, among the 11 epitopes identified, only gp51N11 was capable of inducing CD8(+) T cell-mediated cytotoxicity in all four calves, but it is not a suitable vaccine target because it shows a high degree of polymorphism according to the Wu-Kabat variability index. By contrast, no CTL epitopes were identified from the Gag structural protein. In addition, several epitopes were obtained from gp30 and Tax, indicating that cellular immunity against BLV is strongly targeted to these proteins. CD8(+) CTL epitopes from gp30 and Tax were less polymorphic than epitopes from. Indeed, peptides tax16, tax18, tax19, and tax20 include a leucine-rich activation domain that encompasses a transcriptional activation domain, and the gp30N16 peptide contains a proline-rich region that interacts with a protein tyrosine phosphatase SHP1 to regulate B cell activation. Moreover, at least one CD8(+) CTL epitope derived from gp30 was identified in each of the four calves. These results indicate that BLV gp30 may be the best candidate for the development of a BLV vaccine.

The cathelicidin family is one of the several families of antimicrobial peptides (AMPs). A bovine myeloid antimicrobial peptide (BMAP-28) belongs to this family. Recently, the emergence of drug-resistant bacteria such as methicillin-resistant Staphylococcus aureus (MRSA) has become a big problem. AMPs are expected to be leading compounds of new antibiotics against drug-resistant bacteria. In this study, we focused on the activity of BMAP-28 against bacterial cell surfaces. First, we observed morphological change of MRSA caused by BMAP-28 using a scanning probe microscope. We also studied activities of BMAP-28 against adherence of S. aureus to fibronectin, collagen type I, collagen type IV. We confirmed whether BMAP-28 can bind to lipoteichoic acid (LTA) of S. aureus. BMAP-28 was indicated as damaging the cell surface of MRSA. In a particular range of concentrations, BMAP-28 promoted adherence of S. aureus against fibronectin and collagens. It was revealed that BMAP-28 and LTA of S. aureus bound with each other. Our study showed the potential of BMAP-28 which can damage MRSA and interact with LTA of S. aureus but promote its adherence in some concentrations. This study provides new points of which to take notice when we use AMPs as medicines.

A bovine myeloid antimicrobial peptide antimicrobial peptide (BMAP-28) is a member of the cathelicidin family and acts as a component of innate immunity. There are few reports of susceptibility difference of methicillin-resistant Staphylococcus aureus (MRSA) and susceptible strains (MSSA) against BMAP-28. This study aims to clarify how a few amino acid substitutions of BMAP-28 are related to its antimicrobial activity using four analog peptides of BMAP-28. We also compared cellular fatty acid components of MSSA and MRSA using gas chromatography. We found that a few amino acid substitutions of BMAP-28 do not change antimicrobial activity. It was also revealed that the percentage of cis-11-eicosenoic acid in total detected fatty acids of MRSA was significantly higher than that of MSSA. In addition, the percentage of palmitic acid in total detected fatty acids of MRSA tended to be lower than that of MSSA. Our results will provide new information to deal with the question of differences in bacterial susceptibility against BMAP-28.

The bovine leukemia virus (BLV) is the etiological agent of enzootic bovine leucosis (EBL) which is the most common neoplastic disease of cattle. Recent studies show bovine leukocyte antigen (BoLA)-DRB3 gene plays a direct role in controlling the number of proviral load in cattle and the homozygote BoLA-DRB3*1601 is associated with high proviral load. The other hand, the CD8+ cytotoxic T cell (CTL) response is an important defense against viral invasion. To find the candidate of vaccine for BLV infection we performed CD8+ T cell epitopes mapping in BLV-infected cattle with different BoLA-DRB3 alleles. In this study, 20 amino acids in length of 115 synthetic peptides were made from the sequence of Gag, Env and Tax protein of BLV, overlapping by 10 amino acids. Then total 11 CD8+ T cell epitopes were found out in BLV-infected four Japanese Black cattle with different BoLA-DRB3 alleles that are both of DRB3 *1601/1601, and DRB3 *1501/2703 and DRB3 *1501/0503 allele by WST assay. Different CD8+ T cell epitopes were recognized by cattle with different BoLA-DRB3 alleles and then dose-dependent cytotoxicity activity were measured by non-radioactive cytotoxicity assay. All of four cattle responded to gp51N11, but it has higher variability values by Wu-Kabat variability index analysis. By contrast, CD8+ T cell epitopes were determined from all of four cattle on gp30 region and they have high conservatives. gp30N16 among 4 epitopes was detected in each low proviral load cattle from susceptibility and neutrality groups. Those results were suggested that gp30 region maybe the best candidate for vaccine that can induce cell-mediated immunity against this disease.

Leptospiral lipopolysaccharide (L-LPS) has shown potency in activating toll-like receptor 2 (TLR2) in pig fibroblasts (PEFs_NCC1), and causes the expression of proinflammatory cytokines. However, the stimulation by L-LPS was weak eliciting the function of TLR2 sufficiently in pig innate immunity responses during Leptospira infection. In this study, the immune response of pig embryonic fibroblast cell line (PEFs_SV40) was investigated and was found to be the high immune response, thus TLR2 is the predominate receptor of L-LPS in pig cells. Further, we found a strategy using the antibody against L-LPS, to prevent L-LPS interaction with TLR2 in pig cells which could impact on immune activation.

A bovine myeloid antimicrobial peptide antimicrobial peptide (BMAP-28) is a member of the cathelicidin family and acts as a component of innate immunity. There are few reports of susceptibility difference of methicillin-resistant Staphylococcus aureus (MRSA) and susceptible strains (MSSA) against BMAP-28. This study aims to clarify how a few amino acid substitutions of BMAP-28 are related to its antimicrobial activity using four analog peptides of BMAP-28. We also compared cellular fatty acid components of MSSA and MRSA using gas chromatography. We found that a few amino acid substitutions of BMAP-28 do not change antimicrobial activity. It was also revealed that the percentage of cis-11-eicosenoic acid in total detected fatty acids of MRSA was significantly higher than that of MSSA. In addition, the percentage of palmitic acid in total detected fatty acids of MRSA tended to be lower than that of MSSA. Our results will provide new information to deal with the question of differences in bacterial susceptibility against BMAP-28.

The cathelicidin family is one of the several families of antimicrobial peptides (AMPs). A bovine myeloid antimicrobial peptide (BMAP-28) belongs to this family. Recently, the emergence of drug-resistant bacteria such as methicillin-resistant Staphylococcus aureus (MRSA) has become a big problem. AMPs are expected to be leading compounds of new antibiotics against drug-resistant bacteria. In this study, we focused on the activity of BMAP-28 against bacterial cell surfaces. First, we observed morphological change of MRSA caused by BMAP-28 using a scanning probe microscope. We also studied activities of BMAP-28 against adherence of S. aureus to fibronectin, collagen type I, collagen type IV. We confirmed whether BMAP-28 can bind to lipoteichoic acid (LTA) of S. aureus. BMAP-28 was indicated as damaging the cell surface of MRSA. In a particular range of concentrations, BMAP-28 promoted adherence of S. aureus against fibronectin and collagens. It was revealed that BMAP-28 and LTA of S. aureus bound with each other. Our study showed the potential of BMAP-28 which can damage MRSA and interact with LTA of S. aureus but promote its adherence in some concentrations. This study provides new points of which to take notice when we use AMPs as medicines.

A bovine myeloid antimicrobial peptide (BMAP-28) is a member of the cathelicidin family which is included in the innate immune system of mammals. Recently, there have been many studies about antimicrobial peptides. This study aims to clarify whether BMAP-28 has bactericidal activity against methicillin-resistant Staphylococcus aureus (MRSA) and compares its activity against methicillin-susceptible S. aureus (MSSA) and MRSA. We found that the peptide was effective in killing MRSA (minimal inhibitory concentration (MIC) range; 5-20 µg/mL). It was also revealed that MSSA (MIC range; 1.25-20 µg/mL) had two levels of susceptibility to BMAP-28. We also examined the effect of BMAP-28 on bacterial shape to visually show its activity. After exposure to the peptide, both MSSA and MRSA cells showed the morphological changes on their surfaces. Our results indicate that BMAP-28 is a promising candidate for medicine against drug-resistant bacteria.