B. mori Host Response to Bb Figure 7. Diagram showing the induced biosynthesis, metabolism and signal transduction related genes of juvenile hormone in the silkworm. The expression pattern of each gene was indicated near the gene name. Diagram of the pathway is referenced form Schooley and Baker, 1985. For a detailed view of the cluster ratios, see Table S4. doi:10.1371/journal.pone.0008098.g007 hemocytes including crystocyte, plasmatocyte and lamellocyte, is the first barrier to prevent and remove most viruses and many bacteria, parasites and fungal infections [80]. Many immune effectors, such as lysozyme, lectin, and scavenger receptors (SCRs), also work for the cellular immune response [81]. After Bb enters the silkworm hemolymph, the cellular immune response is triggered (Fig. 8a, Table S4). The expression levels of two lysozymes, which can dissolve and kill bacteria, were modulated. For example, the expression of A007987, a bacterio- phage T7 lysozyme-like protein 1 (BTL-LP1), was upregulated more than three-fold at 24 hpi. The encapsulation process involved in cell adhesion, can lead to bacterial death [80]. Lectins, a type of hemonectin, can condense microorganisms while immune response [82,83]. The condensed microorganisms can then easily be phagocytosed or encapsulated by plasmatocytes and cystocytes, ultimately leading to the formation of black knots. Four of the 21 identified C-type lectin genes were regulated dynamically [23]. Also, C-type lectins (CLTs) participate in signal regulation, whereas the immune signal is transduced during melanization [84]. Of the SCRs, 2 of the 13 identified SCR class B and one class C member were modulated. One class B SCR gene, known to be involved in the phagocytosis of microorganisms in Drosophila, SCRB10, was upregulated at 3 hpi and 24 hpi. In addition, Cu/ Zn superoxide dismutase (SOD1), which can limit parasite development in Anopheles, was also upregulation during the infection [23]. Another SOD gene, SOD2, was upregulated too at 24 hpi. Members of the immunoglobulin superfamily (IgSF), which are related to the specific immune response in vertebrate blood, can also be induced in insects in response to pathogens [85]. After Bb oral infection, the expression levels of 18 IgSF genes were modulated. Boi (A008552), a signal transduction protein, showed upregulation at 24 hpi. Lrig1 (A006920), a leucine-rich repeat immunoglobulin involved in the pathogen response, was also upregulated at 24 hpi [86]. However, two dscam genes (dscam2 and dscam5), known to be involved in cell adherence, were shown to be downregulated in this study [87,88]. Bb induced host serine protease cascade melanization pathway. Melanin formation in the hemolymph is caused by the serine protease cascade melanization pathway after pathogen infection [89–91]. After Bb oral infection has induced an immune response, the serine protease cascade melanization pathway was activated. First, expression of two of the six peptidoglycan recognition protein short-type proteins, PGRP-S2 and PGRP- S5, was induced [23](Fig. 8b). PGRP-S2 (A007987), which is only expressed in the midgut, showed almost five-fold upregulation at 24 hpi. PGRP-S5 (A012866), which is expressed at high levels in the integument and fat body, showed more than three-fold up- regulation at 24 hpi, indicating that Bb can be recognized by PGRP in the fat body and integument [23]. However, none of the six long-type PGRPs showed melanization-related regulation in Drosophila, and four of the b-glucan recognition proteins (bGRP) involved in the PPO-activating system in the silkworm in previous PLoS ONE | www.plosone.org 10 December 2009 | Volume 4 | Issue 12 | e8098